Blended formulations

ABSTRACT

The present invention relates to a formulation comprising blends of formulations (or colloidal dispersions) and its topical application. The formulation comprises at least two different types of colloidal dispersion comprising deformable colloidal particles, wherein the deformable colloidal particles comprise a non-ionic surfactant and/or a phospholipid. The deformable colloidal particles of the invention may comprise an agent of interest (AOI) or may be free of an AOI. The formulation may comprise an AOI that is not associated with the deformable colloidal particles. The present invention also includes kits comprising the formulation of the present invention and the use of the formulation in medicine, skin care and cosmetics.

This application is a continuation of U.S. application Ser. No.15/739,570, filed Dec. 22, 2017, pending, which is the National Stage ofInternational Application No. PCT/EP2016/065415, filed Jun. 30, 2016which claims the benefit of and priority to United Kingdom PatentApplication No. 1511478.8, filed on Jun. 30, 2015, and United KingdomPatent Application No. 1511469.7, filed on Jun. 30, 2015, the contentsof which are incorporated by reference in their entirety.

The present invention relates to a formulation comprising blends offormulations (or colloidal dispersions) and its topical application. Theformulation comprises at least two different types of colloidaldispersion comprising deformable colloidal particles, wherein thedeformable colloidal particles comprise a non-ionic surfactant and/or aphospholipid. The deformable colloidal particles of the invention maycomprise an agent of interest (AOI) or may be free of an AOI. Theformulation may comprise an AOI that is not associated with thedeformable colloidal particles. The present invention also includes kitscomprising the formulation of the present invention and the use of theformulation in medicine, skin care and cosmetics.

Examples of colloidal dispersions that contain colloidal particlescomprising a non-ionic surfactant and a phospholipid, wherein thecolloidal dispersions are free of any pharmaceutically active AOI areknown from WO 2010/140061, which describes the use of drug-free(“empty”) vesicles for the treatment of deep tissue pain, specificallypain associated with osteoarthritis.

Further examples of colloidal dispersions that contain colloidalparticles comprising a non-ionic surfactant and a phospholipid,comprising an AOI are known from WO 2015/014965. WO 2015/014965describes the use of these vesicles for the topical administration of atherapeutic, metabolic or structural AOI that is “tethered” to the lipidand/or the surfactant component of the vesicles, such that the majorityof the AOI is external to the vesicles.

None of these documents disclose or teach a formulation comprisingblends or mixtures of different types of colloidal dispersions, nor dothey disclose or teach such a formulation comprising an AOI that is notassociated with the colloidal particles of the colloidal dispersions.

Citation of any reference in this section of the application is not anadmission that the reference is prior art to the invention. The abovenoted publications are hereby incorporated by reference in theirentirety.

Colloidal dispersions comprising colloidal particles have been used inthe past in attempts to treat skin conditions or as a cosmetic toimprove the appearance of skin. Such colloidal dispersions have alsobeen used to deliver AOIs into the body for the same purposes. In someinstances, such as in WO 2013/171131, an active agent is applieddirectly to the skin and a drug-free colloidal dispersion is appliedover the active agent to “push” it through the skin, it is thought, byway of hydrophilic forces. In other instances, such as in WO2011/022707, drug-free vesicles themselves are used for treating atopiceczema, dishydrotic hand eczema, plaque type psoriasis, seborrheiceczema and acne vulgaris. In WO 2015/014965, examples of colloidaldispersions comprising AOIs are used for different purposes. Forexample, vesicles comprising anti-oxidants or vitamins are used forenhancing the skin's ability to repair and colloidal dispersionscomprising vitamin D may be used to supplement sun cream in order toprevent vitamin D deficiency.

However, these drug-free (i.e. AOI-free) colloidal dispersions andcolloidal dispersions comprising AOIs are applied separately. Therefore,if a user requires the effects of multiple formulations, for example, ifthey have a need to reduce acne-causing bacteria as well as to reducesebum production, they would be required to first apply a colloidaldispersion comprising zinc to kill the bacteria and allow this to drybefore applying a drug-free colloidal dispersion to remove the excesssebum. This results in a number of challenges for the user. For example,where an active agent is applied directly to the skin followed by theapplication of a colloidal dispersion, there is little or no dosecontrol of the application of the active agent. An indeterminate amountof active agent is applied to the skin followed by an indeterminateamount of colloidal dispersion. It is impossible for the user to becertain about how much active agent is being delivered through the skin.In addition, the application of an active agent directly to the skinmeans that it is applied at a high concentration and is more likely tocause adverse skin reactions. Also, if two AOIs are applied without theuse of colloidal dispersions, they can interact, so that the user doesnot know exactly what chemical combination of original products andreaction products they are receiving. Crucially, having to individuallyapply two or more colloidal dispersions is particularly time consumingfor the user as each colloidal dispersion may take up to 10 minutes todry.

The current invention circumvents these problems by combining thedifferent colloidal dispersions into one easy to apply formulation wherethe individual AOIs have not interacted. The colloidal nature of theseblended dispersions allows each dispersion to exist separately andstably until applied to the skin. As such, the colloidal dispersions mayhave a separate function and also be administered together as oneformulation, in one easy application. This saves time for the user,ensures more accurate dosing (on a gram-for-gram basis there is aconsistent ratio of the AOI to the colloidal particles) and reducesadverse skin reactions.

Accordingly, in a first aspect, the present invention provides aformulation comprising a first colloidal dispersion and a secondcolloidal dispersion, wherein each colloidal dispersion comprisesdeformable colloidal particles comprising a surfactant and optionally aphospholipid, and one or more of the deformable colloidal particles ofthe first colloidal dispersion comprise a first agent of interest (AOI).

By “one or more” it is meant that from about 1%, to about 5%, from about5% to about 10%, from about 10% to about 20%, from about 20% to about40%, from about 40% to about 60%, from about 60% to about 80%, fromabout 80% to about 85%, from about 85% to about 90%, from about 90% toabout 92%, to about 94%, to about 96%, to about 97%, to about 98%, toabout 99% or to about 100%, or from about 98% to about 100%.

Where the colloidal dispersions comprise at least one or more deformablecolloidal particles comprising an AOI, it is possible that not all ofthe deformable colloidal particles will comprise an AOI. The AOI isadded during the manufacture of the colloidal dispersions and may tetherto some deformable colloidal particles but not others. For example,depending on the amount of AOI added, there may not be sufficient AOI totether to every colloidal particle.

One or more of the deformable colloidal particles of the secondcolloidal dispersion may also comprise an A01, in which case the AOI ofthe first colloidal dispersion and the AOI of the second colloidaldispersion are different.

One or more of the deformable colloidal particles of the first colloidaldispersion and/or one or more of the deformable colloidal particles ofthe second colloidal dispersion comprise one or more further AOIs andwherein each AOI is different.

The formulation may comprise one or more further colloidal dispersionscomprising deformable colloidal particles comprising a surfactant andoptionally a phospholipid and one or more of the particles of eachdispersion may comprise a further AOI and the first AOI, second AOI andeach further AOI are different. The formulation may include furthercolloidal dispersions comprising deformable colloidal particlescomprising a surfactant and/or a phospholipid with no AOI.

The deformable colloidal particles of the same colloidal dispersion maycomprise more than one type of AOI. For example, the deformablecolloidal particles of the first colloidal dispersion may comprise afirst AOI and a second AOI, wherein the first AOI and second AOI aredifferent. The deformable colloidal particles of the same colloidaldispersion may comprise two, three, four, five or more different typesof AOI. The different types of AOI may all be tethered (attached orbonded) to the colloidal particle within the dispersion in differentcombinations.

The formulation may comprise two or more colloidal dispersionscomprising colloidal particles, wherein the colloidal particles of eachdispersion comprise the same number or different numbers of differenttypes of AOI's. For example, the formulation may comprise a firstcolloidal dispersion that comprises colloidal particles, which have twodifferent types of AOI's attached (bonded or tethered) to them and asecond colloidal dispersion that comprises colloidal particles, whichalso have two different types of AOI's attached to them.

Alternatively, the formulation may comprise a first colloidal dispersionthat comprises colloidal particles, which have two different types ofAOI's attached to them and a second colloidal dispersion that comprisescolloidal particles, which have three different types of AOI attached tothem. The formulation may comprise two or more colloidal dispersions,which comprise colloidal particles that do not have any AOI tethered tothem or that have one, two, three, four, five or more different types ofAOI tethered to them.

The colloidal dispersions are also known as “Sequessome™, Transfersome™or Tethersome intermediates”.

The deformable colloidal particles of the invention may be vesicles. TheAOI may be incorporated into the vesicle membrane. The AOI may be bondedto a component of the vesicle membrane. More specifically, the AOI maybe bonded to a surfactant component of the vesicle membrane or to alipid component of the vesicle membrane. Both a surfactant component anda lipid component of the vesicle membrane may have an AOI bonded tothem. The bond may be a covalent bond.

The AOI may be bonded to a component of the particle such that at leasta portion of the AOI is on the external surface of the particle, and isexternal to the particle membrane. Preferably, the component to whichthe AOI is bonded is a lipid and/or a surfactant component. At least aportion means that of the total AOI that is external to the particle atleast 5%, 10% or 20%, suitably 40%, or more than 50% of each molecule(in terms of size or volume of the molecule) is external to the membraneof the particle. Preferably, the majority of the AOI, more preferablythe entire AOI molecule is external to the particle. The AOI may becovalently bonded to a component such that it presents wholly on theexternal surface of the particle.

By the AOI being “external to the particle”, it is meant those AOI thatare “facing outwards”. During one method of manufacture of theparticles, whereby the surfactant or lipid component that is bonded orotherwise attached to the AOI is mixed with the unmodified components,the orientation of the modified molecule cannot be controlled. Thus,approximately 50% of the molecules to which the AOI is attached will bein the “incorrect” orientation, meaning that a portion of the AOI willbe present in the lumen of the particle.

During another method of manufacture of the particles, whereby thesurfactant or lipid component that is bonded or otherwise attached tothe AOI is mixed with already formed particles, the surfactant or lipidcomponent attached to the AOI will preferentially combine with theexisting particle such that AOI is usually left external to theparticle. Thus approximately 0% to 10% of the molecules to which the AOIis attached will be in the “incorrect” orientation, meaning that a smallportion if any of the AOI will be present in the lumen of the particle.Of the modified molecules that are in the “correct” orientation suchthat the AOI is external to the particle, at least 50% of the AOImolecule itself, in terms of physical size/volume, is external to thevesicular membrane. The manufacturing process may result in a lowerproportion of the AOI being external to the particle i.e. the externalconcentration of the AOI may be between 1% to 10% (including 2%, 3%, 4%,5% , 6% , 7% 8% or 9%) 10% to 50%, 15% to 45%, 20% to 40% or 25% to 30%(wt/vol) of the total AOI in the formulation. By external concentrationit is meant the concentration of AOI that is available for releaseand/or to exert its therapeutic activity once the particles havepenetrated the skin.

A particle of the formulation may have a single or a plurality of AOIsbonded to its external surface. Wherein a plurality of AOIs are bonded,the AOIs may all be the same, i.e. homogenous, or the AOIs may bedifferent, i.e. heterogeneous.

The AOI to be bonded may be covalently or otherwise bonded directly toeither the phospholipid or surfactant component of the particle or thelipid component of the particle. It may be desirable to use a link orbridge that is covalently or otherwise bonded to both the fatty acid,surfactant or lipid component and the AOI. In one example, if aninorganic AOI were to be added (for example a metal, a metal salt or ametal oxide), an additional linker, for example a metal chelating agent(such as EDTA), stearic acid or palmitic acid might first be conjugatedto the particle component. In another example it may be desirous to usea longer molecule, for example a polymer (such as polyethylene glycol;PEG), to facilitate the efficacy of the bonding process and/or theeffectiveness of the bound AOI. Such linkers/longer bridging moleculeswill be particularly of benefit when it is desirable to hold an AOI atsuch a distance from the particles in order to prevent it interferingwith the membrane itself. This may occur if the AOI is particularlylipophilic.

The AOI may be bonded (or attached or tethered) to the surfactantcomponent of the colloidal particle. The bonding to the surfactant maybe directly onto the surfactant by ester bond if the molecule has ahydroxyl group. An alternative method of bonding is to substitute anatom or functional group of the surfactant (for example in the case ofTween, a polyethylene glycol polymer) with the AOI. A third method ofbonding is directly to a fatty acid, optionally via an ester bond. Ifthe AOI is an inorganic molecule then a further linking molecule canfirst be conjugated to the vesicle component, for example a metalchelating agent such as EDTA in the case of a metal salt. If it isdesirous that the AOI be held at some distance from the vesicle in orderto maximise its efficiency (for example to expose an active site on anadditional ingredient that is an enzyme), then a linking molecule, forexample a polymer chain (for example polyethylene glycol) may be bondedto both a component of the colloidal particle and the AOI.

The AOI may be attached (bonded or tethered) to the lipid component ofthe colloidal particle. The bonding to the lipid might be achieved viaany of the glycerol hydroxyl groups by an ester bond, for example byeliminating a fatty acid and replacing with the AOI. Alternatively, themethod of attachment may be by replacement of the phosphatidyl moietysuch that the final molecule has two fatty acid chains together with thetethered AOI. The modified lipid inserts in the aliphatic region asnormal and with the free rotation available on the glycerol template,the tethered AOI would locate on the outside of the vesicle. An amidebond may be used for a more stable alternative, should the AOI berequired to be tethered to the deformable colloidal particle for alonger duration. This may be desirable, for example, if the target forthe AOI is deeper tissue, such as the reticular layer of the dermis,rather than the upper dermal layers (e.g. the epidermis). A combinationof less stable and more stable bonds may be used (e.g. ester and amide,respectively) to achieve staggered release of the AOI.

The method of bonding to any component may be hydrolysable ornon-hydrolysable. If it is desirable that the AOI should be detachedonce within or under the skin, the link should be hydrolysable. If it isdesirable that the bonded AOI should remain bound to the vesicle oncewithin or under the skin, the link should be non-hydrolysable.

The AOI may be covalently bonded or conjugated to a membrane component;the bond may be hydrophilic or hydrophobic or hydrostatic; the bond maybe a hydrogen bond or an ionic bond.

The terms “bonding”, “attaching” and “tethering” are used hereinthroughout interchangeably to encompass all the bonds mentioned above.

The AOI may be selected from the group consisting of an element, apeptide, a protein, a micronutrient, an ion, an inorganic salt, a smallmolecule, an amino acid, a carbohydrate, a lipid, a macromolecule or amacrocyclic molecule.

The AOI may be a skin structural protein (such as elastin or collagen),a therapeutic protein, porphyrin or chromophore containing amacromolecule, a vitamin (such as vitamin C, D or E) titanium dioxide,zinc, zinc oxide, zinc stearate, melanin or a melanin analogue. The AOImay be a peptide or synthetic organic chemical such as ananti-inflammatory drug, such as an NSAID.

The first AOI, the second AOI and/or each further AOI may be selectedfrom the group consisting of zinc, ascorbate (vitamin C), vitamin D,tocopherol (vitamin E), tetrapeptide, tripeptide, salicylic acid ormenthol. Specifically, the AOI may be selected from the group consistingof zinc stearate, palmitoyl ascorbate (or palmitoyl ascorbic acid(PAA)), palmitoyl tripeptide, palmitoyl tetrapeptide, salicylic acid ormenthol. More specifically, the AOI may be selected from the groupconsisting of zinc stearate, palmitoyl ascorbate, palmitoyltripeptide-1, palmitoyl tetrapeptide-7, tocopherol linoleate, salicylicacid or menthol. The salicylic acid may be myristyl salicylate ortridecyl salicylate.

Peptides, such as tetrapeptide-7 or tripeptide-1, may be bonded to afatty acid or surfactant component of the vesicle membrane.Tetrapeptide-7 may be useful in fighting inflammation and act tostimulate skin regeneration by way of collagen production. This meansthat it is particularly useful in skin care, and anti-ageing products.Tripeptide-1 has a similar action. Efficient delivery through theexternal layer of skin may provide increased effects at lowerlevels/concentrations thus minimising possible side effects resultingfrom the suppression of interleukins.

Vitamin C (water-soluble ascorbate) is a cofactor in many enzymaticreactions including several collagen synthesis reactions. Thesereactions are important in wound healing and in preventing bleeding fromcapillaries. Therefore, the current invention includes associatingascorbate with a vesicle component, for incorporation both into skincareand suncare preparations to ameliorate damage to collagen, and forincorporation into wound care products.

Vitamin E has important anti-oxidant properties and is able to reducethe signs of ageing and sun damage. Linoleate is the ester of linoleicacid, an omega-6 fatty acid, which has anti-inflammatory andmoisturisation properties. Tocopheryl linoleate combines the propertiesof both vitamin E and linoleate to reduce the signs of ageing and tomoisturise the skin. Therefore, vitamin E or tocopheryl linoleate may beassociated with a vesicle component for incorporation into anti-ageingproducts and skin repair products.

Vitamin D may be incorporated into formulations in accordance with thepresent invention by association with a vesicle component.

Mammalian skin makes vitamin D₃ through the action of UV radiation onits precursor, 7-dehydrocholesterol, and supplies about 90 percent ofour vitamin D. Sunscreen absorbs ultraviolet light and prevents it fromreaching the skin.

When the formulation comprises vitamin D₃ (which is not to say that italso does not comprise vitamin C and/or E) the formulation may beincorporated into a sunscreen product, a sun block product, an after-sunproduct or other skincare or cosmetic product to supplement low vitaminD levels. Low vitamin D levels may be caused by low light conditions, oruse of sunblock, which can prevent the manufacture of vitamin D withinthe body.

Zinc is an antioxidant and antimicrobial. It is beneficial for theimmune system and helps to prevent ageing, to speed up wound healing, tokill microbes and bacteria and to down-regulate sebum production bysebaceous glands. It is currently used in a wide range of topical skinpreparations, such as in sun creams to protect against sunburn and inanti-dandruff shampoo to kill dandruff causing fungus. It is also usedin the treatment of acne. In the present invention, zinc is tethered tovesicle components for incorporation into skin care products that can,for example, be applied to red, dry, itchy skin, to red, irritated,scaly skin or to acne prone skin in order to improve these conditions.

Salicylic acid is a beta hydroxy acid (BHA) with the formulaC₆H₄(OH)COOH. It can be obtained from the bark of willow trees.Salicylic acid is an important active metabolite of aspirin(acetylsalicylic acid), which acts in part as a pro-drug to salicylicacid. It is widely used as an analgesic and it also hasanti-inflammatory, anti-pyretic, anti-diabetic, bactericidal andantiseptic effects. As a result, it is an important ingredient inpain-killers, topical anti-acne products, rubefacient products,skin-care products for the treatment of conditions such as psoriasis,calluses, ichthyosis and warts, shampoos for the treatment of dandruff,suntan and sunscreen products, mouthwashes and dentifrices. It can alsobe used for the prevention or treatment of uneven skin tone caused by,for example, darker melanic spots and liver spots. Salicylic acid can beused in the form of myristyl salicylate or tridecyl salicylate.

A plurality of AOIs may be bonded to a lipid or surfactant component topresent on the external surface of the particle so that once takenthrough the skin, they continue to present on the surface of theparticle. Examples include anti-oxidants; vitamins; inorganic compoundssuch as TiO₂ and ZnO; porphyrin molecules for use in photodynamictherapies or any of the AOIs mentioned above.

As will be appreciated, other AOIs may be tethered to the deformablecolloidal particles of the invention in order to treat a wide variety ofdiseases.

The formulation may or may not contain any known therapeutic agent,other than the AOI bonded to the particles. The formulation comprisingan AOI may or may not be free of any further AOI. A further AOI may ormay not be associated with the colloidal particles.

The AOI may be an agent of interest, such as a cosmetic, that does nothave any biological or pharmacological effect or it may be abiologically active or pharmaceutically active agent.

The colloidal dispersions of the formulation comprise at least twophases; a dispersed phase comprising the suspended deformable colloidalparticles and a continuous phase, comprising the medium of suspension(or matrix). The formulation may comprise two, three, four or moredispersed phases. The dispersed phases may comprise different types ofcolloidal particle such as different types of vesicle, for exampleSequessomes™, Transfersomes™, Tethersomes, micelles and liposomes. Theformulation may comprise one or more dispersed phases comprisingdeformable colloidal particles such as Sequessomes™, Transfersomes™ orTethersomes and optionally one or more dispersed phases comprisingnon-deformable colloidal particles such as micelles and/ornon-deformable vesicles such as liposomes. The formulation may comprisetwo or more types of deformable colloidal particle.

Usually, the term “Transfersome™” is used to refer to a deformablephospholipid and surfactant vesicle, which is associated with an AOI, inparticular where the AOI is held either in the lumen of the vesicle orbound within the vesicle's membrane. As described herein, the term“Sequessome™” is usually used to refer to the deformable phospholipidand surfactant vesicle itself. If an AOI is ‘tethered’ to the vesicle,such that it is held outside the external surface of the membrane, thevesicle (or Sequessome™) may be referred to as a ‘Tethersome’.

The AOI may be present in the continuous phase. Preferably, thiscontinuous phase comprises a non-lipid phase, most preferably an aqueousphase.

Alternatively, the AOI may be present in a second dispersed phase (thefirst dispersed phase comprising the deformable colloidal particles).For example, the AOI may be associated with non-deformable colloidalparticles such as micelles or non-deformable vesicles such as liposomes.

The formulation may be a liquid, cream, lotion, ointment, gel, solution,spray, lacquer or film forming solution. Preferably, the formulation maybe a gel.

As mentioned above, the formulation of the first aspect of the inventionmay comprise an AOI which is not associated with the deformablecolloidal particles in the formulation.

As used herein, the phrase “not associated with” means that the AOI isnot attached to or contained within the deformable colloidal particle.In particular, the AOI is not tethered, bound or otherwise secured tothe deformable colloidal particle, is not contained within the structureof the deformable colloidal particle, including the lipid bilayer or thefluid enclosed by the lipid bilayer, is not encapsulated within thedeformable colloidal particle or in any other way directly associatedwith the deformable colloidal particle. Rather, the AOI is present in adifferent phase of the composition from the deformable colloidalparticles. For example, the

AOI may be present within the continuous phase of the composition, forexample dissolved in the continuous phase. Alternatively, the AOI may bepresent within a different dispersed phase of the composition, forexample in the form of insoluble aggregates or associated withnon-deformable colloidal particles such as micelles and non-deformablevesicles such as liposomes. Preferably, the AOI is in a form in which itwould not normally pass through the skin, without the assistance ofdeformable colloidal particles.

The formulation of the present invention is multiphasic because, asmentioned above, it comprises at least two phases; a dispersed phasecomprising the suspended deformable colloidal particles and a continuousphase comprising the medium of suspension. In a multiphasic system thereare discrete particles, assemblies or bodies dispersed or suspendedwithin a medium or matrix. Each phase is a separate solid or liquidentity with a detectable phase boundary. In colloidal systems, theparticle size of each phase is too small for observation with the nakedeye. However, the multiphasic nature of the system can be demonstratedby applying a narrow beam of light, a Tyndall Beam, the passage of whichis visible through the solution due to scattering of the light by thephase boundary(ies). The passage of such a beam of light through asingle phase solution would not be visible.

The formulation of the present invention may comprise more than twophases. For example, the formulation may comprise two, three, four ormore dispersed phases, in addition to the continuous phase. Two suchdispersed phases may comprise the deformable colloidal particles whichdrive the AOI through the skin. A third dispersed phase may comprise theAOI, for example in the form of insoluble aggregates or associated withnon-deformable colloidal particles. Other dispersed phases may also beprovided comprising further deformable and/or non-deformable colloidalparticles.

The formulation may comprise two or more colloidal dispersions.

Preferably, the continuous phase comprises a non-lipid phase. Thecontinuous phase may be selected depending upon the identity of the AOIand whether the AOI is to be dissolved in the continuous phase orprovided as a dispersed phase. Preferably, the continuous phase is anaqueous phase.

Whilst an AOI may be provided in a separate phase from the deformablecolloidal particles, under certain circumstances, which will be wellknown to those of skill in the art, some of the AOI may partition intothe membrane or the fluid core of the deformable colloidal particles.Preferably, only a negligible or insignificant amount of the AOIprovided in a separate phase from the deformable colloidal particlespartitions into the deformable colloidal particles whilst a significantamount of the AOI remains in a separate phase. Preferably, about 5% orless, about 4% or less, about 3% or less, about 2% or less, about 1% orless, about 0.5% or less, about 0.1% or less or about 0% of the AOIpartitions into the deformable colloidal particles. Preferably, at leastabout 95.0%, at least about 96.0%, at least about 97.0%, at least about98%, at least about 99.0%, at least about 99.5%, at least about 99.9% orabout 100% of the AOI in the formulation remains in a separate phasefrom the deformable colloidal particles.

The formulation according to the first aspect of the present inventionmay comprise two significant populations of the same AOI; a firstpopulation that is not associated with the deformable colloidalparticles and a second population that is associated with the deformablecolloidal particles, for example, tethered to the deformable colloidalparticles. In such embodiments, the first population of the AOI willusually penetrate less deeply into the skin than the second populationof the AOI, providing effects at different depths of tissue.

Where the formulation of the first aspect of the present inventioncomprises two such populations of the same AOI, the proportion of thefirst population to the second population is designed according to theseparate purposes that each of the populations is designated to perform.Preferably, the ratio of the first population to the second populationis within the range 20:80 to 80:20, 30:70 to 70:30, 40:60 to 60:40 orapproximately 50:50.

As used herein, the phrase “Agent of Interest” or “AOI” includes but isnot limited to biologically active or pharmaceutically active agents. Abiologically or pharmaceutically active agent is herein defined as anagent that has pharmacological, metabolic or immunological activity.This may include nutraceuticals or pharmaceuticals. Where necessary, theAOI has preferably received regulatory approval.

The AOI may be an antiseptic, an antibiotic, an anaesthetic, ananalgesic, a skin lightener, an antihistamine, a steroid, ananti-inflammatory agent, an anti-viral, sun block, moisturiser,nicotine, an anti-fungal, an antimicrobial, a nutraceutical, anessential oil or a hormone.

Preferably, the AOI that is not associated with the deformable colloidalparticles is selected from the group including but not limited tochlorhexidine or a salt thereof, capsaicin, salicylic acid or a salt orester thereof, glucosamine or a salt thereof, an amide of glucosamine ora salt thereof, chondroitin or a salt thereof, caffeine or tocopherol.

If the AOIs listed above are added to the organic phase of theformulation during manufacture (i.e. prior to formation of thedeformable colloidal particles), some AOI may be present inside thelumen of the colloidal particles and some AOI may be present in thecontinuous phase of the formulation (i.e. not associated with thecolloidal particles). The distribution of the AOI may depend on theparticular nature of the AOI that is included in the formulation.

Chlorhexidine,N,N″″1,6-Hexanediylbis[N′-(4-chlorophenyl)(imidodicarbonimidicdiamide)], is a cationic polybiguanide (bisbiguanide). It is anantimicrobial compound that is effective against a wide range ofgram-positive and gram-negative bacteria and some fungi and viruses. Itis widely used in topical disinfectants, cosmetics and pharmaceuticalproducts, including wound dressings and mouthwashes. It is also used forthe treatment of acne.

Chlorhexidine may be frequently used in the form of a salt. Preferably,the salt of chlorhexidine is selected from the group consisting ofchlorhexidine dihydrochloride, chlorhexidine diacetate, chlorhexidinegluconate and chlorhexidine digluconate. Most preferably, theformulation comprises chlorhexidine gluconate.

The chlorhexidine ora salt thereof of the present invention may beprovided as a 10% solution, a 20% solution, a 30% solution, a 40%solution or a 50% solution. Preferably, chlorhexidine or a salt thereofmay be provided as a 20% solution. The formulation of the presentinvention may comprise 0.1 to 2.5%, more preferably 0.5 to 2.0%, mostpreferably 1.5% by weight of the 10% solution, 20% solution, 30%solution, 40% solution or 50% solution. Preferably, the formulation ofthe present invention may comprise 0.1 to 2.5%, more preferably 0.5 to2.0%, most preferably 1.5% by weight of the 20% solution. Theformulation may therefore comprise from 0.02% to 0.1%, from 0.1% to0.2%, from 0.2% to 0.3%, from 0.3% to 0.4%, from 0.4% to 0.5%, from 0.5%to 1.1% or from 1.1% to 1.25% of chlorhexidine. Preferably, theformulations of the present invention comprise chlorhexidine or a saltthereof at a concentration below its critical micelle concentration.

The chlorhexidine or salt thereof may be present in the formulation inthe form of insoluble aggregates or micelles. Where the AOI notassociated with the deformable colloidal particles compriseschlorhexidine or a salt thereof, the continuous phase is preferablyaqueous.

Capsaicin, 8-methyl-N-vanillyl-6-nonenamide, is a capsaicinoid which isproduced as a secondary metabolite by the fruits of plants in the genusCapsicum, including chili peppers. It is believed to act as a deterrentagainst predation by certain mammals and also as an anti-fungal agent.In medicine, capsaicin is used as an analgesic, particularly for thetemporary relief of pain associated with arthritis, backache, strainsand sprains, fibromyalgia and neuralgia. It is also used to reduceitching associated with conditions such as psoriasis.

The formulations of the present invention may comprise 0.01 to 0.1%capsaicin, more preferably 0.01 to 0.05%, most preferably approximately0.025%.

The capsaicin may be dissolved in the continuous phase, when thecontinuous phase comprises a suitable solvent, for example ether,benzene and/or an alkane.

Preferably, the capsaicin is present in the formulation in the form ofinsoluble aggregates or associated with non-deformable colloidalparticles such as liposomes comprising one or more phospholipids.Preferably, the capsaicin is bound in the liposome membrane. Under suchcircumstances, the continuous phase is preferably aqueous.

As mentioned above, salicylic acid is widely used as an analgesic and italso has anti-inflammatory, anti-pyretic, anti-diabetic, bactericidaland antiseptic effects.

Salicylic acid can be used in the above applications in the form of asalt or ester. Salts of salicylic acid include calcium salicylate,magnesium salicyalte, MEA-salicylate, potassium salicylate, sodiumsalicylate and TEA-salicylate. Esters of salicylic acid includebutyloctyl salicylate, C12-15 alkyl salicylate, capryolyl salicylicacid, hexyldecyl salicylate, isocetyl salicylate, isodecyl salicylate,ethylhexyl salicylate, methyl salicylate, myristyl salicylate andtridecyl salicylate.

Preferably, the formulation of the first aspect of the present inventioncomprises an ester of salicylic acid, most preferably myristylsalicylate or tridecyl salicylate.

The composition of the present invention may comprise salicylic acid ora salt or ester thereof in concentrations appropriate to the intendeduse and within any regulatory limitations.

Salicylic acid and salts and esters thereof are water soluble and thus,where the continuous phase is aqueous, the salicylic acid or a salt orester thereof may be dissolved within the continuous phase. At leastsome of the salicylic acid or salt or ester thereof may also partitioninto the membrane of the deformable colloidal particles of the presentinvention and/or into the membrane of one or more non-deformablecolloidal particles also present in the formulation. The formulations ofthe present invention preferably comprise about 0.05% to 2.5% by weightof salicylic acid or a salt or ester thereof.

In solution, chlorhexidine and salts thereof, capsaicin, and salicylicacid and esters and salts thereof may form micelles, vesicles orinsoluble aggregates.

Glucosamine, or (3R,4R,5S)-3-Amino-6-(hydroxymethyl)oxane-2,4,5-triol,is an amino sugar and a prominent precursor in the biochemical synthesisof glycosylated proteins and lipids. It is commonly used as a dietarysupplement, in particular for the treatment of osteoarthritis.

Glucosamine may be used in the form of an amide such asN-acetylglucosamine.

The formulations of the present invention may comprise salts ofglucosamine or the amides of glucosamine, such as glucosamine sulfate,glucosamine hydrochloride and N-acetylglucosamine sulphate.

The formulations of the present invention may comprise about 0.01 to1.0% by weight of glucosamine, amides of glucosamine or salts thereof,more preferably about 0.05 to 0.5% by weight, more preferably about 0.1to 0.3% by weight, most preferably approximately 0.2% by weight.

Glucosamine, amides of glucosamine and salts thereof are water soluble.Thus, where the continuous phase of the formulation is aqueous, thesemay be dissolved within the continuous phase.

Chondroitin is a glycosaminoglycan (GAG) composed of a chain ofalternating sugars, N-acetylgalactosamine and glucuronic acid. It isusually found attached to proteins as part of a proteoglycan. Achondroitin chain can have over 100 individual sugars, each of which canbe sulfated in variable positions and quantities.

Chondroitin may be used in the form of a salt, such as chondroitinsulphate, chondroitin gluconate and chondroitin hydrochloride.Chondroitin sulfate is an important structural component of cartilageand has become a widely used dietary supplement for treatment ofosteoarthritis.

The formulations of the present invention may comprise about 0.01 to1.0% chondroitin or salts thereof by weight, more preferably about 0.05to 0.5% by weight, more preferably about 0.1 to 0.3% by weight, mostpreferably approximately 0.2% by weight.

Chondroitin and salts thereof, such as chondroitin sulphate, are watersoluble. Thus, where the continuous phase of the formulation is aqueous,these may be dissolved within the continuous phase.

Caffeine, or 1,3,7-Trimethylpurine-2,6-dione, is a methylxanthinealkaloid which is naturally found in the seeds, nuts, or leaves of anumber of plants native to South America and East Asia. Whilst commonlyused as a stimulant of the central nervous system, caffeine is also apowerful antioxidant and anti-inflammatory that can be topically appliedto reduce wrinkles, eye puffiness and dark under-eye circles. It mayalso be used to prevent sun damage to the skin.

The formulations of the present invention may comprise about 0.001 to1.0% caffeine by weight, more preferably about 0.01 to 0.5% by weight,more preferably about 0.02 to 0.1% by weight, more preferably about 0.02to 0.08% by weight, most preferably approximately 0.05% by weight.

Caffeine is water soluble. Thus, where the continuous phase of theformulation is aqueous, caffeine may be dissolved within the continuousphase. At least some of the caffeine may also partition into themembrane of the deformable colloidal particles of the present inventionand/or one or more non-deformable colloidal particles also present inthe formulation.

The formulations of the present invention may comprise more than one AOIthat is not associated with the deformable colloidal particles. Forexample, a formulation of the present invention may comprise glucosamineor a salt thereof and/or an amide of glucosamine or a salt thereof, incombination with chondroitin or a salt thereof. For example, theformulation of the present invention may comprise glucosaminehydrochloride and/or N-acetylglucosamine sulphate in combination withchondroitin sulphate.

Tocopherols are a class of organic chemical compounds (more precisely,various methylated phenols), many of which have vitamin E activity.There are four different forms of tocopherol; alpha, beta, gamma anddelta. Tocopherols have a number of functions within the body. They haveantioxidant and anti-inflammatory effects, which help protect cells fromthe damaging effects of free radicals which can lead to cardivasculardisease and cancer.

Tocopherols are involved in healthy immune function, gene expression,blood circulation, protecting the health of nerves and preventing mentaldegeneration. Tocopherols also have an important role in skin care,protecting the skin from oxidative damage resulting from UV rays andpollution. Topical application of tocopherols can help reduce redness,sunburn and skin damage, including UV-induced tumour formation.Tocopherols also help to protect the cells that make collagen andelastin, providing an anti-ageing effect. Tocopherols are also aneffective moisturiser, hydrating the skin and preventing further waterloss, aiding in the reduction of fine lines and wrinkles. Tocopherolsalso reduce the healing time of wounds and can be used to help repairskin lesions and dry skin and treat skin conditions such as psoriasisand erythema.

Alpha-tocopherol is the form of vitamin E that is preferentiallyabsorbed and accumulated in humans.

Tocopherol, preferably alpha-tocopherol, may be present in thecontinuous phase of the compositions of the present invention (as “free”tocopherol).

The compositions of the present invention may comprise about 0.01 to1.0% by weight of “free” tocopherol, such as alpha tocopherol, morepreferably 0.05 to 0.5% by weight, more preferably 0.1% to 0.3% byweight, most preferably approximately 0.2% by weight. A small amount ofthe “free” tocopherol may partition into the lumen of the deformablecolloidal particles, but a significant amount will remain in thecontinuous phase.

Exemplary compositions according to the first aspect of the presentinvention comprising alpha tocopherol in the continuous phase areprovided in Example 6 and Example 7.

Derivatives of tocopherols, for example esters such as tocopheryllinoleate, may also be used. Such derivatives may be tethered to thedeformable colloidal particles in various compositions of the presentinvention, as discussed herein.

The formulation may comprise a least three colloidal dispersions, atleast four colloidal dispersions or at least five or more colloidaldispersions.

The formulation of the first aspect of the invention comprises asurfactant and optionally a phospholipid from which the deformablecolloidal particles are formed. The deformable colloidal particles maycomprise one or more surfactants and no phospholipids, one or morephospholipids and no surfactant, or one or more surfactants incombination with one or more phospholipids.

Preferably, these deformable colloidal particles comprise a fluid core,for example an aqueous core, enclosed by a membrane, for example abilayer membrane. Suitable bilayer membranes includephospholipid-surfactant bilayer membranes, non-ionic surfactant bilayermembranes and surfactant-cholesterol bilayer membranes. The term“bilayer” indicates that the membrane is two molecules in depth, whereineach molecule is arranged with its hydrophobic end directed inwardstowards the opposite side of the membrane and its hydrophilic enddirected outwards towards the surrounding aqueous media. For example,the bilayer membrane may comprise two layers of phosphatidylcholinemolecules, wherein the hydrophobic tails of each molecule point towardeach other and the hydrophilic heads of each molecule point away fromeach other. Preferably, the formulation comprises deformable colloidaldispersions comprising colloidal particles (i.e. vesicles) having alipid bilayer in aqueous media. As used herein, the term “vesicle”includes unilamellar or multilamellar vesicles and micelles. Thedeformable colloidal particles may also comprise niosomes.

Preferably, these deformable colloidal particles are approximately 50 nmto 250 nm in diameter, approximately 60 nm to 200 nm in diameter,approximately 70 nm to 180 nm in diameter, approximately 80 nm to 160 nmin diameter, preferably 100 to 150 nm in diameter, most preferablyapproximately 120 nm in diameter.

Deformable colloidal particles of this invention are described in bothWO 2010/140061 and in WO 2011/022707.

The formulation of the present invention is particularly suitable fortopical administration as it comprises ultra-deformable (or flexible)colloidal particles (or microscopic spheres). These flexible forms ofcolloidal particles (“Sequessomes™, Transfersomes™ or Tethersomes”) arevesicles, which are bilayer vesicles composed of surfactant and/orlipid, such as phospholipid. The specific amount of surfactant (whenpresent), preferably non-ionic surfactant, modifies the lipid membraneto such an extent that the resulting particles are in a permanent liquidcrystalline state. Since the surfactant (when present) also confersmembrane stability, the particles are also ultra-deformable and stable(have reduced rigidity without breaking).

The colloidal dispersions of the formulation form into deformablecolloid particles suspended in a suspension media, for example, anaqueous buffer. The particles are highly hydrophilic and this property,together with their ultra-deformability, is key to their ability to betransported across the skin. When the formulation of the invention isapplied to the skin and allowed to dry, the rehydration driving force ofthe particles combined with their deformability gives rise to movementof the particles to areas of higher water content on and below the skinpermeability barrier. This drives their movement through skin pores andintracellular gaps. It may be the specific ratio of lipid to surfactantthat facilitates transdermal delivery of the particles. As the colloidalparticles move through the skin, they push or pull the AOIs with them.

While not wishing to be limited to any mechanism of action, theformulations of the invention are able to form colloidal particlescharacterized by their deformability. The movement of the deformablecolloidal particles through the pores and intracellular gaps carrieswith it the AOI, increasing the speed, depth and effectiveness ofabsorption of this compound.

The term “deformable” refers to the ability of the colloidal particlesto easily change their properties, such as shape, elongation ratio andsurface to volume ratio. The colloidal particles of this invention maybe characterized by their ability to adjust their shape and propertiesto the anisotropic stress caused by crossing narrow pores in asemipermeable barrier such as the skin. Sufficient deformability impliesthat a colloidal particle can sustain different unidirectional forces orstress, such as one caused by pressure, without extensive fragmentation,which defines a “stable” colloidal particle.

A “barrier” in the context of this invention is a body withthrough-extending narrow pores, such narrow pores having a radius whichis at least 25% smaller than the radius of the colloidal particles(considered as spherical) before said colloidal particles permeatethrough such pores.

The term “narrow” used in connection with a pore implies that the poreradius is significantly, typically at least 25%, smaller than the radiusof the colloidal particle tested with regard to its ability to cross thepore. The necessary difference typically should be greater for thenarrower pores. Using a 25% limit is therefore quite suitable for >150nm diameter, whereas >100% difference requirement is more appropriatefor the smaller systems, e.g., with <50 nm diameter.

Preferably, the deformability of the colloidal particles may bedetermined by the ability of the colloidal particles to penetrate abarrier with pores having an average pore diameter at least 25%, atleast 30%, at least 35%, at least 40%, at least 55%, at least 50%, atleast 55% or at least 60% smaller than the average particle diameterbefore the penetration. Most preferably, the deformability of thecolloidal particles may be determined by the ability of the colloidalparticles to penetrate a barrier with pores having an average porediameter at least 50% smaller than the average particle diameter beforethe penetration.

The term “semipermeable” used in connection with a barrier implies thata solution can cross transbarrier openings whereas a suspension ofnon-adaptable aggregates (large enough for the above definition of“narrow” pores to apply) cannot. Conventional lipid vesicles (liposomes)made from any common phosphatidylcholine in the gel lamellar phase orelse from any biological phosphatidylcholine/cholesterol 1/1 mol/molmixture or else comparably large oil droplets, all having the specifiedrelative diameter, are three examples for such non-adaptable aggregates.

The term “stable” means that the colloidal particles do not change theirdiameter spontaneously or under the transport related mechanical stress(e.g. during passage through a semipermeable barrier) unacceptably,which most often means only to a pharmaceutically acceptable degree. A20-40% change is normally considered acceptable; the halving or doublingof the diameter of the colloidal particles is borderline and a greaterchange in diameter is typically unacceptable. Alternatively and veryconveniently, the change in diameter of the colloidal particlesresulting from pore crossing under pressure is used to assess systemstability; the same criteria are then applied as for “narrow” pores,mutatis mutandis. To obtain the correct value for diameter change, acorrection for flux/vortex effects may be necessary. These proceduresare described in greater detail in the publications of the applicant inCevc et. al., Biochim. Biophys. Acta 2002; 1564:21-30.

Non-destructing passage of colloidal particles through narrow pores in asemi-permeable barrier is thus diagnostic of deformability. If poreradius is two times smaller than the average radius of the colloidalparticles, the colloidal particles must change their shape andsurface-to-volume ratio at least 100% to pass without fragmentationthrough the barrier. An easy and reversible change in shape inevitablyimplies high deformability of the colloidal particles and requires largesurface-to-volume ratio adaptation. A change in surface-to-volume ratioper se implies: a) high volume compressibility, e.g. in the case ofcompact droplets containing material other than, and immiscible with,the suspending fluid; b) high membrane permeability, e.g. in the case ofcolloidal particles that are free to exchange fluid between inner andouter vesicle volume.

Methods of testing deformability which may be used to characterise thecomposition of the invention are set forth in WO 2010/140061 and USPatent Application Nos. 2004/0071767 and 2004/0105881, each hereinincorporated by reference as if set forth herein in their entirety.

Where a formulation in accordance with the present invention comprisestwo or more colloidal dispersions comprising deformable colloidalparticles comprising an AOI and, in some cases, an AOI not associatedwith the colloidal particles, the formulation is able to exert itseffects in different layers of the skin. The colloidal particles areable to efficiently penetrate the skin and deliver the AOI to deeperlayers of the dermis, whilst the AOI agent may remain on the surface ofthe skin or in the upper layers. This ensures that each area of the skincan be targeted in a way that most effectively treats the specificcondition.

In a second aspect, the present invention provides a formulationcomprising a first colloidal dispersion and a second colloidaldispersion, wherein each colloidal dispersion comprises deformablecolloidal particles comprising a surfactant and optionally aphospholipid and one or more of the deformable colloidal particles ofthe first colloidal dispersion comprise a first agent of interest (AOI),for use in treating or preventing a disease.

In a third aspect, the present invention provides a formulationcomprising a first colloidal dispersion and a second colloidaldispersion, wherein each colloidal dispersion comprises deformablecolloidal particles comprising a surfactant and optionally aphospholipid and one or more of the deformable colloidal particles ofthe first colloidal dispersion comprise a first agent of interest (AOI),for cosmetic treatment of a subject or for use in skin care.

The formulation may be used for treating or preventing acne. Theformulation may be used for treating or preventing dry skin, itchy skin,red skin, irritated skin, scaly skin, ageing skin, thinning skin,periorbital skin, uneven skin tone, photo-aged skin or dermatitis. Theformulation may also be used for cosmetic treatment of these conditionsi.e. to improve the appearance of skin affected by these conditions.

Treating or preventing periorbital skin may be the treatment orprevention of wrinkles around eyes (i.e. around the eye sack), finelines around eyes, dark circles around eyes, eye bags, sagging skinaround eyes, hyperpigmentation around eyes or puffiness around eyes.

The use for which the formulation of the present invention is providedwill depend at least in part on the identity of the first AOI and/or thesecond AOI and/or any further AOI included in the formulation.

For example, where a formulation in accordance with the first aspect ofthe invention comprises a first drug-free colloidal dispersion, a secondcolloidal dispersion comprising deformable colloidal particles tetheredto zinc stearate and a further AOI that is chlorhexidine or a saltthereof, the formulation may be used for treating or preventing acne ordermatitis. This formulation can also be used to treat acne-prone skin,in particular to prevent the appearance of acne in acne-prone skin.

Acne is a common skin condition that affects many people at some pointin their lives. It causes spots to develop on the skin, usually on theface, back and chest. The spots can range from surface blackheads andwhiteheads, which are often mild, to deep, inflamed, pus-filled pustulesand cysts, which can be severe and lead to scarring. The condition iscaused by over-production of sebum which in turn results in increasedand changed bacterial activity leading to inflammation and pus.

Current treatment options for acne range from simple cleansers totopical exfoliators and antibiotics, to oral retinoid preparations,which can be associated with serious side effects. Many patients eitherfind the treatments ineffective or limited by side effects. Theformulation of the present invention circumvents these problems.

The formulation may be useful for the treatment or prevention of acne,not only because the deformable colloidal particles increase the speed,depth and effectiveness of absorption of the zinc stearate and thechlorhexidine or salt thereof into the skin of the patient, but alsobecause the colloidal particles of the first drug-free colloidaldispersion can facilitate the clearance of lipids from the skin byfacilitating sebum removal. Furthermore, as discussed above, tetheringzinc stearate to the deformable colloidal particles down-regulates sebumproduction by sebaceous glands and including chlorhexidine in theformulation kills bacteria and controls bacterial hyper-proliferation.As such, this formulation is able to clear existing sebum and reduce itsproduction by denying microbes of their ‘nutrient source’, as well askilling exist microbes. These multiple functions help to calm inflamedspots, support epidermal renewal, remove impurities and tighten andconstrict pores, resulting in a reduction in symptom recurrence (such asspots, hormonal blemishes, blackheads and oily skin) and further spreadof infection.

Where a formulation in accordance with the first aspect of the inventioncomprises a first drug-free colloidal dispersion and a second colloidaldispersion comprising deformable colloidal particles tethered to zincstearate, the formulation may be used for treating or preventing red,irritated, scaly skin. The colloidal particles of the first colloidaldispersion in this formulation sequester sebum and the zinc stearateattached to the colloidal particles of the second colloidal dispersionreduces sebum production. This dual function reduces colonisation ofsecretions by microbes, maintaining the normal skin environment and alsoreduces reappearance of symptoms by denying microbes their ‘nutrientsource’; the sebum on which they grow. In addition, the colloidalparticles deliver nutrients to the skin. As such, this formulationreduces redness, swelling, flaking, weeping, blistering and skin erosioncaused by excess oil and also promotes healthy skin balance and supportsthe skin's own renewal and repair mechanism.

For the avoidance of doubt, references herein to “drug-free” or “empty”are to colloidal dispersions that do not contain any non-lipidnon-surfactant AOI that has a therapeutic purpose. These colloidaldispersions may comprise active agents such as antimicrobials,stabilisers and preservatives. However, it is to be understood thatthese agents are simply for improving the stability of the formulationsand for increasing their shelf-life; they do not have a therapeuticpurpose.

Where a formulation in accordance with the first aspect of the inventioncomprises a first drug-free colloidal dispersion, a second colloidaldispersion comprising deformable colloidal particles comprisingpalmitoyl ascorbate, a third colloidal dispersion comprising deformablecolloidal particles comprising palmitoyl tetrapeptide-7 and a fourthcolloidal dispersion comprising deformable colloidal particlescomprising palmitoyl tripeptide-1, the formulation may be used fortreating or preventing ageing and thinning skin.

The symptoms associated with ageing and thinning skin are welldocumented and publicised. Equally, there are numerous cosmetictreatments available that target such conditions but many of thesetreatments are not supported by robust data. In addition, each of thesecosmetic treatments has to be applied separately to target each symptomof ageing, thinning skin. The formulations of the present inventionsimplify this and ensure that all of the symptoms of ageing can beeffectively treated in one application of the formulation.

Palmitoyl ascorbate is a form of vitamin C that acts as antioxidant;shielding skin from free radical damage and reducing the appearance ofbrown spots from sun damage, boosting healthy collagen production andreducing inflammation and irritation. Palmitoyl tetrapeptide-7 andPalmitoyl tripeptide-1 both supress the production of excessinterleukins and thus inhibit unnecessary inflammatory responses thatlead to glycation damage and stiffening of tissues. In addition, thesepeptides boost the growth of connective tissues, naturally increasingthe production of collagen in the skin and allowing the skin to heal andrejuvenate itself. Zinc stearate and tocopherol linoleate may beoptionally added to enhance the performance of this blend: zinc isimportant in slowing the breakdown of collagen and tocopherol (vitaminE) is an important anti-oxidant that can stabilise vitamin C and enhanceits effect. Therefore, this formulation can address all visible signs ofskin photo-ageing, ensuring a more lifted appearance, better definitionand noticeably firmer skin. It helps to build new collagen and elastin,slows collagen breakdown and strengthens the skin's underlyingstructure. It also preserves the extracellular dermal matrix, preventsfree radical damage and hydrates and restores. It reduces fine lines,wrinkles, eye bags, dark circles, skin sagging and hyperpigmentation andimproves skin tone, smoothness, firmness, elasticity, complexion.

Where a formulation in accordance with the first aspect of the inventioncomprises a first drug-free colloidal dispersion, a second colloidaldispersion comprising deformable colloidal particles comprisingpalmitoyl ascorbate, a third colloidal dispersion comprising deformablecolloidal particles comprising palmitoyl tetrapeptide-7, a fourthcolloidal dispersion comprising deformable colloidal particlescomprising palmitoyl tripeptide-1 and a fifth colloidal dispersioncomprising deformable colloidal particles comprising zinc stearate, theformulation may be used for treating or preventing red, dry, itchy skin.Red, dry, itchy skin may be due to conditions such as atopic eczema anddishydrotic hand eczema. As mentioned above, palmitoyl ascorbate worksas an antioxidant to prevent skin damage, peptides promote collagenproduction to enable skin regeneration and zinc acts as an antimicrobialand reduces sebum production and also retards collagen breakdown. Thecolloidal particles themselves ensure deep dermal hydration with purewater and prevent water loss by forming an intimate evaporation barrierwithin the skin structure. As such, this formulation functions to defendthe skin, to support the skin's own repair mechanism and to delivernutrients to the skin to improve natural skin function, helping to avoidnew flare-ups. This reduces redness, swelling, flaking, weeping,itching, skin thickening, blistering and skin erosion, which are oftensymptoms of skin conditions such as eczema.

Where a formulation in accordance with the first aspect of the inventioncomprises a first colloidal dispersion comprising deformable colloidalparticles comprising tridecyl salicylate and alpha tocopherol orderivatives thereof and a second colloidal dispersion comprisingcolloidal particles comprising palmitoyl ascorbic acid, tocopheryllinoleate and alpha tocopherol or derivatives thereof, the formulationmay be used for treating or preventing uneven skin tone. The alphatocopherol in this formulation may be present within the lumen of thecolloidal particles or in the continuous phase of the formulation (i.e.not associated with the colloidal particles). The tridecyl salicylate,palmitoyl ascorbic acid and tocopherol linoleate may be tethered to thecolloidal particle. The formulation may improve the appearance of unevenskin tone in one or more of the following ways; reducing the appearanceof wrinkles, hyperpigmentation, blemishes, dark under-eye circles,improving the appearance of skin texture and increasing skin elasticity.

This formulation may also be used for treating or preventing pain ordiscomfort by acting as a topical counter-irritant.

Where a formulation in accordance with the first aspect of the inventioncomprises an AOI that is selected from glucosamine or a salt thereof, anamide of glucosamine or a salt thereof, and/or chondroitin or a saltthereof, the formulation may be used for maintaining and prolongingjoint health and/or for treating or preventing joint diseases(arthropathy), in particular degenerative joint diseases such asarthritis and osteoarthritis. These formulations may also be used totreat or prevent the pain associated with poor joint health.

Where a formulation in accordance with the first aspect of the inventioncomprises an AOI that is selected from glucosamine or a salt thereof, anamide of glucosamine or a salt thereof, and/or chondroitin or a saltthereof, the formulation may also be used for the manufacture of amedicament for maintaining and prolonging joint health and/or fortreating or preventing joint diseases (arthropathy), in particulardegenerative joint diseases such as arthritis and osteoarthritis, or thepain associated with poor joint health.

Where a formulation in accordance with the first aspect of the inventioncomprises an AOI that is selected from glucosamine or a salt thereof, anamide of glucosamine or a salt thereof, and/or chondroitin or a saltthereof, the AOI may or may not be associated with (i.e. bonded to) thedeformable colloidal particle. If the AOI is associated with thedeformable colloidal particle, a derivative a glucosamine or a saltthereof, an amide of glucosamine or a salt thereof, and/or chondroitinor a salt thereof may be used to allowing tethering (i.e. bonding) tothe deformable colloidal particle.

The formulations of the present invention may be useful for maintainingand prolonging joint health and for treating or preventing jointdiseases or the pain associated with poor joint health, not only becausethe deformable colloidal particles increase the speed, depth andeffectiveness of penetration of the AOI into the joint of the patient,but also because the deformable colloidal particles can themselvesalleviate or attenuate pain, such as that associated withosteoarthritis, as discussed above.

The volumetric blending ratio (by weight) of the individual colloidaldispersions (or Sequessome™, Transfersome™ or Tethersome intermediates)may differ in each formulation depending on the specific type ofcolloidal dispersion. For example, in the formulation that may be usedto treat ageing thinning skin, the ratio of the palmitoyl ascorbateintermediate to the tetrapeptide intermediate and the tripeptideintermediate may be from about 1:1:1 to about 2:1:1, from about 1:2:1 toabout 1:3:1, from about 1:1:2 to about 1:1:3, preferably from about3:1:1 to about 5:1:1 and most preferably about 3:1:1. In the formulationthat may be used to treat red, dry, itchy skin, the ratio of thepalmitoyl ascorbate intermediate to the tetrapeptide intermediate, thetripeptide intermediate and the zinc stearate intermediate may be fromabout 1:1:1:1 to about 2:1:1:2, from about 3:1:1:3 to about 3:1:1:4,from about 4:2:2:5 to about 5:3:3:7, preferably from about 3:1:1:5 toabout 4:2:2:6 and most preferably about 3:1:1:5. In the formulationsthat may be used to treat red, irritated, scaly skin and acne prone skinthe ratio of the drug-free intermediate to the zinc stearateintermediate may be from about 1:1 to about 2:1, from about 3:1 to about3:2, from about 3:2 to about 4:3 and most preferably from about 1:1.

Where the formulation comprises capsaicin as the AOI not associated withthe deformable colloidal particles, the formulation may be used fortreating or preventing one or more conditions selected from the groupconsisting of pain, including muscle pain, joint pain and itching.Capsaicin stimulates the nerve endings in the skin to create a warmingsensation (without actually being related to a physical temperaturechange). As such, capsaicin containing formulations can be applied tothe muscle or joint or any other tissue to relieve aches or pains.Formulations comprising capsaicin can be used prophylactically, beingapplied prior to exercise to create a warming sensation in the skin andunderlying muscles and joints and thus prevent muscle soreness. Theformulations may also be applied during or post-exercise to relieveaches and pains. Formulations of the invention comprising capsaicin arealso provided for use as an analgesic medicament or an antipruriticmedicament.

Where the formulation comprises salicylic acid or a salt or esterthereof, which is associated with and/or not associated with thecolloidal particles, the formulation may also be used for treating orpreventing one or more conditions selected from the group consisting ofpain, including muscle pain and joint pain. Salicylate is an analgesicand has anti-inflammatory properties. As such, salicylate containingformulations can be applied to the muscle or joint or any other tissueto relieve aches or pains. Formulations comprising salicylate can beused prophylactically, being applied prior to exercise to relieve painin the skin and underlying muscles and joints. The formulations may alsobe applied during or post-exercise to relieve aches and pains and reduceinflammation in the muscle and joint tissue.

The formulations comprising both capsaicin and salicylic acid or a saltor ester thereof may be used for treating or preventing one or moreconditions selected from the group consisting of pain, including musclepain, joint pain and itching. These formulations may have an enhancedeffect in treating pain, such as muscle pain and joint pain, as they areable to both warm the skin and underlying muscles and joints and alsoprovide an analgesic and anti-inflammatory effect in these areas.

The formulations comprising capsaicin and/or salicylic acid may beuseful for the treatment or prevention of pain, not only because thedeformable colloidal particles increase the speed, depth andeffectiveness of absorption of the capsaicin and/or salicylic acid intothe skin of the patient, but also because the colloidal particles canthemselves alleviate or attenuate pain, such as that associated withosteoarthritis, as discussed above.

Formulations of the present invention are particularly suitable forpatients with chronic, long-term pain associated with joint diseasessuch as osteoarthritis, who wish to avoid or minimise the consumption ofpharmaceutical painkillers.

Where the AOI comprises glucosamine or a salt thereof, an amide ofglucosamine or a salt thereof, and/or chondroitin or a salt thereof, thedisease, disorder or condition is preferably a joint disease(arthropathy), in particular a degenerative joint disease such asarthritis or osteoarthritis, or the pain associated with poor jointhealth.

The present invention also provides a method of maintaining andprolonging joint health, comprising topically applying a formulationaccording to the first aspect of the invention to the skin of a patientin need thereof, wherein the AOI comprises glucosamine or a saltthereof, an amide of glucosamine or a salt thereof, and/or chondroitinor a salt thereof. The AOI may or may not be associated with (i.e.bonded to) the deformable colloidal particles. When associated,derivatives of glucosamine and/or chondroitin will be used.

Formulations of the present invention comprising capsaicin haveapplications in cosmetics through their mild irritant effect. Inparticular, the topical application of capsaicin to the skin results invasodilation. When applied to the lips, capsaicin causes the lips toswell and redden, enhancing their appearance.

Formulations of the present invention comprising salicylic acid or asalt or ester thereof and tocopherol can be used to improve theappearance of uneven skin tone by reducing the appearance ofhyperpigmentation such as melanic spots and liver spots. The salicylicacid or a salt or ester thereof and the tocopherol may be present in theformulation, both associated with the colloidal particles and notassociated with the colloidal particles. The salicylic acid or salt orester thereof and the tocopherol which is associated with the colloidalparticles may be tethered to the colloidal particles. The tetheredsalicylic acid may be in the form of myristyl salicylate or tridecylsalicylate, in which the myristyl aliphatic chain or tridecyl aliphaticchain is embedded in the colloidal particles. The tethered tocopherolmay be in the form of tocopheryl linoleate, in which the linoleate chainis embedded in the colloidal particles.

Formulations of the present invention may comprise colloidal dispersionscomprising colloidal particles comprising myristyl salicylate ortridecyl salicylate, wherein the myristyl salicylate or tridecylsalicylate is tethered to a component of the colloidal particle.

Formulations of the present invention may comprise colloidal dispersionscomprising colloidal particles comprising tocopheryl linoleate, whereinthe tocopheryl linoleate is tethered to a component of the colloidalparticle.

These formulations may not comprise any other form of salicylic acid ortocopherol. These formulations may comprise one or more furthercolloidal dispersions comprising colloidal particles comprising adifferent AOI, such as vitamin C. Alternatively, the colloidal particlescomprising myristyl salicylate, tridecyl salicylate or tocopheryllinoleate may also comprise vitamin C. The formulations may alsocomprise tocopherol that is not associated with the colloidal particles(i.e. it may be present in the continuous phase of the formulations).The formulations may also comprise tocopherol that is present in thelumen of the vesicles.

Formulations of the present invention may comprise colloidal dispersionscomprising drug-free colloidal particles and myristyl salicylate ortridecyl salicylate, wherein the myristyl salicylate or tridecylsalicylate is not associated with the colloidal dispersions. In theseformulations, myristyl salicylate or tridecyl salicylate may formmicelles in the media, which may be a separate phase to the colloidalparticles.

Such a formulation has a dual effect. The tethered salicylic acid orsalt or ester thereof will penetrate more deeply into the stratum basaleof the skin (where the melanocytes are located) where it will exert ananti-tyrosinase action to down-regulate melanin production. Thesalicylic acid or salt or ester thereof which is not associated with thedeformable colloidal particles will penetrate less deeply into the skin,remaining in the outer layers of the epidermis where it will exert amild exfoliating effect, which will increase the speed of turnover ofnew, now de-melanised, skin. The skin-lightening effects of the tetheredsalicylic acid or salt or ester thereof will therefore be seen by thepatient more quickly than in the absence of untethered salicylic acid ora salt or ester thereof.

A product containing a salicylate compound may find use as a topicalcounter-irritant.

Formulations of the present invention comprising caffeine can improvethe appearance of the skin, particularly periorbital skin. As discussedabove, they do so by acting as a powerful antioxidant andanti-inflammatory. In particular, caffeine has vasoconstrictorproperties and is thought to shrink the blood vessels that causeunder-eye dark circles. As demonstrated in Example 6, the formulationsof the present invention can reduce the appearance of puffiness andswelling around the eyes and below the lower eyelid (eye bags),under-eye dark circles and fine lines and deep wrinkles, includingcrow's feet, around the eye area. The formulations can also result inperiorbital skin which looks and feels less thin, looks and feels morefirm, feels more elastic, looks smoother and which is more hydrated. Theformulations can also lift sagged skin, resulting in a reduction in skindroopiness and sagging. The formulations can make the skin tone lookmore even, make the eye contour look and feel tighter and look moretoned and lifted and make the eyes look rested and less tired. They mayalso be used to prevent sun damage to the skin.

Without wishing to be bound by any particular theory, it is alsobelieved that the colloidal particles themselves have a beneficialeffect on the skin, particularly periorbital skin, as they have a‘nano-stenting’ effect that helps this sensitive tissue to drain excessfluid. By ‘nano-stenting’ effect it is meant that the colloidalparticles act as a form of proppant or scaffold and locate in theextracellular spaces of tissues where they facilitate the draining ofextracellular/interstitial fluids into the lymph. It is believed thatthe colloidal particles physically push the interstitial fluid andby-products contained therein. This assists in the removal ofundesirable by-products to the lymph and through the lymph, preventingtoxic build-up, and also assists in the normal drainage of the lymphwhere this has been compromised.

Where these formulations comprise palmitoyl ascorbic acid andtocopherol, their antioxidant effect is enhanced. Where theseformulations comprise palmitoyl tripeptide-1 and palmitoyltetrapeptide-7 tethered to the deformable colloidal particles, collagensynthesis is also stimulated. The appearance of the periorbital skin istherefore further improved.

Compositions of the present invention comprising tocopherol in thecontinuous phase and salicylic acid or a salt or ester thereof tetheredto the deformable colloidal particles can improve the appearance of theskin. In particular, the compositions can be used to improve theappearance of skin tone, for example by treating and/or preventinguneven skin tone.

As discussed above, the tocopherol acts as a powerful antioxidant andanti-inflammatory, protecting the cells that make collagen and elastinand acting as an effective moisturiser. As discussed above, salicylicacid or a salt or ester thereof can also improve the appearance ofuneven skin tone by reducing the appearance of hyperpigmentation such asmelanic spots and liver spots. As demonstrated in Example 7, thecompositions of the present invention can reduce the appearance of finelines and wrinkles and the amount and size ofhyperpigmentation/pigmented skin blemishes. The compositions can resultin skin which looks significantly smoother and feels significantly morehealthy and elastic. The compositions can help the complexion lookyounger and significantly healthier, be visibly improved and moreradiant. The compositions can also make skin tone look significantlymore even and significantly lighten hyperpigmentation/pigmented skinblemishes and periorbital dark circles.

Where these compositions comprise palmityol ascorbic acid and tocopheryllinoleate tethered to a second form of deformable colloidal particle,their antioxidant effect is enhanced.

The compositions of the present invention may be useful for improvingthe appearance of the skin, not only because the deformable colloidalparticles increase the speed, depth and effectiveness of absorption ofthe tocopherol and salicylic acid or salt or ester thereof into the skinof the patient, but also because the deformable colloidal particlesthemselves can improve the appearance of the skin. As discussed above,the deformable colloidal particles can enhance drainage of theinterstitial fluid to the lymph nodes, preventing toxic build-up ofby-products in the skin.

The invention also provides the use of a formulation according to thefirst aspect of the present invention for the manufacture of amedicament for the treatment or prevention of a disease or for cosmetictreatment of a subject or for skin care.

For all of the formulations in accordance with the invention, it is theparticular combination (or blend or mixture) of the specific colloidaldispersions and, when present, the AOI that allows the formulation tohave multiple effects.

Formulations in accordance with the present invention may be known as a“dermocosmetics” as they are able to address the symptoms ofdermatological conditions as well as mask them. Without wishing to bebound by any theory, it is the flexible microscopic spheres (i.e. thecolloidal particles) of the formulation of the present invention,containing naturally occurring plant lipids, that enables theformulation to supplement the skin, support the natural recoveryprocess, ensure deep dermal hydration with pure water and prevent waterloss by forming an intimate evaporation barrier within the skinstructure. When present, the

AOI in the continuous phase of the formulation (i.e. that is notassociated with a deformable colloidal particle) is pushed through theskin by the deformable colloidal particles in the formulation and anyAOI that is associated with a deformable colloidal particle is pulledthrough the skin by the deformable colloidal particles to which it isattached. These actions ensure that the AOIs penetrate the skin and aredelivered in effective amounts to the required areas.

The formulations of the present invention may comprise one or morevitamins. These vitamins may be tethered to the colloidal particles,present in the continuous phase or as a separate phase.

Such vitamins may include vitamin C (ascorbic acid) or esters thereof,for example palmitoyl ascorbic acid or palmitoyl ascorbate. Theformulations may comprise 0.01 to 1.0% by weight vitamin C (ascorbicacid) or esters thereof, more preferably 0.05 to 0.5% by weight, mostpreferably approximately 0.1% by weight. Preferably, the vitamin C oresters thereof are tethered to the deformable colloidal particles.

Another preferred vitamin for inclusion in formulations of the presentinvention is vitamin E (tocopherol) or esters thereof, for exampletocopheryl linoleate. The formulations may comprise about 0.01 to 1.0%by weight vitamin E (tocopherol) or esters thereof, more preferablyabout 0.05 to 0.5% by weight, most preferably about 0.1% to 0.2% byweight. Preferably, the tocopheryl linoleate is tethered to thedeformable colloidal particles. Preferably, tocopherol is present as aseparate phase.

Vitamins such as vitamin C and vitamin E and esters thereof provide thevesicles with an anti-oxidant protective effect.

Formulations of the present invention may comprise both vitamin C andvitamin E or esters thereof. Preferably, the vitamin C or esters thereofand the vitamin E or esters thereof, where present, are tethered toseparate vesicles.

The tethering of such vitamins is particularly preferred when the AOIwhich is not associated with the deformable colloidal particlescomprises salicylic acid or a salt or ester thereof, glucosamine or asalt thereof, an amide of glucosamine or a salt thereof, chondroitin ora salt thereof, caffeine or tocopherol.

Preferably, when the formulation comprises salicylic acid or a salt orester thereof, palmitoyl ascorbic acid and tocopheryl linoleate aretethered to the deformable colloidal particles and tocopherol is presentas a separate phase. In a particularly preferred embodiment, theformulation comprises two forms of deformable colloidal particle; thefirst comprising salicylic acid or a salt or ester thereof tetheredthereto and the second comprising palmitoyl ascorbic acid and tocopheryllinoleate tethered thereto.

Preferably, when the formulation comprises glucosamine or a saltthereof, an amide of glucosamine or a salt thereof and/or chondroitin ora salt thereof, the formulation additionally comprises palmitoylascorbate and tocopheryl linoleate tethered to the deformable colloidalparticles. The inclusion of such vitamins into these formulationsenhances the efficacy of these formulations in maintaining andprolonging joint health and treating or preventing joint diseases or thepain associated with poor joint health.

Preferably, when the formulation comprises caffeine, palmitoyl ascorbicacid and tocopheryl linoleate are tethered to the deformable colloidalparticles and tocopherol is present as a separate phase.

Where the formulations contain tocopherol, a small amount of tocopherolmay be present within the lumen of the deformable colloidal particles,with the majority of tocopherol present within the continuous phase ofthe formulations.

Formulations of the present invention may additionally comprisetripeptide-1, preferably palmitoyl tripeptide-1, and/or tetrapeptide-7,preferably palmitoyl tetrapeptide-7, tethered to deformable colloidalparticles.

The formulations may comprise about 0.0001 to 0.1% by weight oftripeptide-1, preferably palmitoyl tripeptide-1, more preferably about0.001 to 0.01% by weight, more preferably about 0.002 to 0.008% byweight, most preferably approximately 0.006% by weight.

The formulations may comprise about 0.0001 to 0.01% by weight oftetrapeptide-7, preferably palmitoyl tetrapeptide-7, more preferablyabout 0.001 to 0.01% by weight, more preferably about 0.002 to 0.008% byweight, most preferably approximately 0.006% by weight.

The tethering of tripeptide-1 and tetrapeptide-7, in particularpalmitoyl tripeptide-1 and/or palmitoyl tetrapeptide-7, to deformablecolloidal particles is particularly preferred when an AOI which is notassociated with the deformable colloidal particles is caffeine. In aparticularly preferred embodiment, the formulation comprises three typesof deformable colloidal particle; the first comprising palmitoylascorbic acid tethered thereto and tocopherol, the second comprisingpalmitoyl tripeptide-1 tethered thereto and the third comprisingpalmitoyl tetrapeptide-7 tethered thereto. The tocopherol may be presentin the lumen of the colloidal particles, or outside the colloidalparticles in the continuous phase of the formulation (i.e. notassociated with the colloidal particles).

The inclusion of palmitoyl tripeptide-1 and palmitoyl tetrapeptide-7into these formulations enhances their efficacy by stimulating collagensynthesis, firming the skin and reducing the appearance of wrinkles.

The formulation of the invention may be used to treat any animal. Theanimal to be treated may be a human, a companion animal (e.g. a cat, adog or a horse) or an agricultural animal. When the animal to be treatedis a human, the invention may be useful for the treatment of patients ofany age, for example, patients aged from 15 years old to 100 years old.The human patient may be male or female.

As discussed above, the formulation may be topically applied, enablingthe formulation of the invention to be particularly useful as a cosmeticor for treating skin conditions.

The invention comprises a formulation for use as herein described,wherein any amount of the formulation may be topically applied as oftenas desired, provided this is within any regulatory restrictions imposedon the use of the AOI in the formulations. For example, the formulationmay be applied once, twice, three times or more per day. Alternativelythe formulation may be administered on alternate days, two or threetimes per week, once per week or less frequently as needed. Theformulation may be administered over a period of one or more weeks, forexample, for at least one week, two weeks, three weeks, four weeks, fiveweeks, six weeks, seven weeks, eight weeks, nine weeks, ten weeks,eleven weeks, or twelve weeks, sixteen weeks, twenty four weeks, fourmonths, six months, eight months, ten months, one year, two or moreyears, or indefinitely as needed.

Any amount of the formulation sufficient to treat any of the conditionsdisclosed herein may be administered to the patient. For example, a 0.1to 10 gram dose of the formulation of the invention may be administeredto the patient. The dose may be 1 to 10 grams, or 1 to 5 grams or about1 gram, 2 grams, 3 grams, 4 grams, 5 grams, 6 grams, 7 grams, 8 grams, 9grams or 10 grams. The dose may be measured as the total weight of theformulation. The dose can be measured as the total weight of thephospholipid(s) and/or surfactant(s) in the formulation. The dose may beadministered once or twice daily or as often as needed. The dose may beadministered once, twice, three, four, five, six, or seven times perweek, or as often as needed, in accordance with the invention. The dosemay be administered every day, every other day, or two to three times aweek, or as often as needed, in accordance with the invention.Preferably, the formulation may be applied twice daily. A suitableamount of the formulation (i.e. any amount of the formulation sufficientto treat a condition as herein described) may be spread over the problemarea of the skin. The formulation may be left to dry for up to 15minutes.

Where the formulation is to be used as a cosmetic, any amount of theformulation may be topically applied to the skin as often as necessaryto achieve the desired effect.

The lipid in the deformable colloidal particles of the formulation maybe phosphatidylcholine.

The surfactant in the deformable colloidal particles of the formulationmay be polysorbate 80.

The formulation may include one or more buffers, chelators, humectants,lubricants, antioxidants, preservatives, microbicides, antimicrobials,emollients, co-solvents or thickeners. The thickeners may be Carbopol®(polyacrylamide) or methylcellulose.

In a fourth aspect, the present invention provides a method ofcosmetically improving the appearance of a subject comprisingadministering the formulation of the present invention to the subject.Preferably, the formulation is topically applied to the skin of thesubject, who is preferably human.

Where the AOI comprises chlorhexidine or a salt thereof, the methodpreferably comprises improving the appearance of the skin of thesubject, for example by treating or preventing acne.

Where the AOI comprises capsaicin, the method preferably comprisesimproving the cosmetic appearance of the lips of the subject, forexample by plumping the lips.

Where the AOI agent comprises salicylic acid or a salt or ester thereofand/or tocopherol or a derivative thereof, the method preferablycomprises improving the appearance of the skin, for example by treatingor preventing uneven skin tone, preferably by reducing the appearance ofhyperpigmentation.

Where the AOI comprises caffeine, the method preferably comprisesimproving the appearance of the skin of the subject, preferably theperiorbital skin.

Where the AOI comprises glucosamine or salt thereof, an amide ofglucosamine or a salt thereof and/or chondroitin or a salt thereof, themethod preferably comprises improving the appearance of the skin, forexample by enhancing collagen production to reduce the appearance offine lines and wrinkles.

In a fifth aspect, the present invention provides a method of making thecomposition of the first aspect of the invention. Preferably, the methodcomprises a primary manufacturing step in which a colloidal dispersionis made from an “organic phase” containing alcohol-soluble componentsand an “aqueous phase” consisting of water-soluble components. During asecondary manufacturing step, this initial dispersion is mixed with athickener to form a gel with the desired consistency. The AOI ispreferably added during this secondary manufacturing step. More than onekind of colloidal dispersion may be introduced such that the finalcomposition comprises more than one kind of colloidal particle.

In a sixth aspect, the present invention provides a method of treating adisease, disorder or condition in a subject comprising topicallyapplying a formulation comprising a first colloidal dispersion and asecond colloidal dispersion, wherein each colloidal dispersion comprisesdeformable colloidal particles comprising a surfactant and optionally aphospholipid and one or more of the deformable colloidal particles ofthe first colloidal dispersion comprise a first agent of interest (AOI)and wherein the disease is treated.

Where the AOI comprises chlorhexidine or a salt thereof, the disease,disorder or condition is preferably acne or dermatitis.

Where the AOI comprises capsaicin, the disease, disorder or condition ispreferably selected from the group comprising pain, including musclepain, and itching.

Where the AOI agent comprises salicylic acid or a salt or ester thereofand/or tocopherol or a derivative thereof, the disease, disorder orcondition is preferably pain or discomfort.

Where the AOI comprises glucosamine or a salt thereof, an amide ofglucosamine or a salt thereof, and/or chondroitin or a salt thereof, thedisease, disorder or condition is preferably a joint disease(arthropathy), in particular a degenerative joint disease such asarthritis or osteoarthritis, or the pain associated with poor jointhealth.

The present invention also provides a method of maintaining andprolonging joint health, comprising topically applying a compositionaccording to the first aspect of the invention to the skin of a patientin need thereof, wherein the AOI comprises glucosamine or a saltthereof, an amide of glucosamine or a salt thereof, and/or chondroitinor a salt thereof.

The present invention can be used to administer the AOI, such aschlorhexidine or a salt thereof, capsaicin, salicylic acid or a salt orester thereof, glucosamine or salt thereof, an amide of glucosamine or asalt thereof and/or chondroitin or a salt thereof, caffeine ortocopherol, to or through the skin of an animal. Preferably, the methodcomprises topically applying the composition to the skin of the patientin an amount sufficient to penetrate the skin to deliver the AOI.

The present invention can also be used to administer an AOI, such asglucosamine or a salt thereof, an amide of glucosamine or a salt thereofand/or chondroitin or a salt thereof, to a joint of an animal, forexample a shoulder, elbow, wrist, knuckle, knee or ankle joint.

Preferably, the method comprises topically applying the composition tothe skin of the patient in an amount sufficient to penetrate the skin todeliver the AOI to the underlying joint. Any animal can be included,including humans, dogs, cats, horses, food production animals and pets.

In a seventh aspect, the present invention provides a kit comprising oneor more compartments, wherein at least one compartment contains aformulation comprising a first colloidal dispersion and a secondcolloidal dispersion, wherein each colloidal dispersion comprisesdeformable colloidal particles comprising a surfactant and optionally aphospholipid and one or more of the deformable colloidal particles ofthe first colloidal dispersion comprise a first agent of interest (AOI).

Preferably, the formulation according to the first aspect of theinvention is contained within a single compartment of said kit.

The kit may be formatted such that it is marked to indicate quantity ofthe formulation remaining or dispensed.

The kit may be in the form of a tube, sachet or pot. The kit may alsocomprise a dispensing means, preferably selected from group consistingof a pump, nozzle, measuring cup or spatula.

The instructions for administration of the formulation according to thefirst aspect of the invention preferably comprise instructions for theadministration of the formulation in accordance with any of the second,third, fourth, fifth or sixth aspects of the present invention.

Preferably, the instructions comprise instructions for the topicalapplication of the formulation.

The kit may be for use in the treatment of disease.

Where the AOI comprises chlorhexidine or a salt thereof, theinstructions for administration thereof preferably comprise instructionsfor administration thereof to a patient or subject in need thereof forthe treatment or prevention of a disease or disorder associated with theskin of a patient, preferably acne or dermatitis. The instructions mayalternatively direct the subject to administer the composition to theskin for the purpose of improving its appearance.

Where the AOI comprises capsaicin, the instructions for administrationthereof preferably comprise instructions for administration thereof to apatient or subject in need thereof for the treatment or prevention ofpain, including muscle pain, and/or itching. The instructions may alsodirect the subject to administer the composition prior to exercise. Theinstructions may alternatively direct the subject to administer thecomposition to the lips for the purpose of providing a cosmetic“plumped” effect.

Where the AOI agent comprises salicylic acid or a salt or ester thereofand/or tocopherol or a derivative thereof, the instructions foradministration thereof preferably comprise instructions foradministration thereof to a patient or subject in need thereof for thetreatment or prevention of pain or discomfort. The instructions mayalternatively direct the subject to administer the composition to theskin for the purpose of improving the appearance of uneven skin tone,including hyperpigmentation.

Where the AOI comprises glucosamine or salt thereof, an amide ofglucosamine or a salt thereof, and/or chondroitin or a salt thereof, theinstructions for administration thereof preferably comprise instructionsfor administration thereof to a patient or subject in need thereof formaintaining and prolonging joint health and/or for treating orpreventing joint diseases (arthropathy), in particular degenerativejoint diseases such as arthritis and osteoarthritis, or the painassociated with poor joint health. The instructions may alternativelydirect the subject to administer the composition to the skin for thepurpose of improving the appearance of the skin, for example by reducingthe appearance of fine lines and wrinkles and improving the appearanceskin tone, barrier function and hyperpigmentation.

Where the AOI comprises caffeine, the instructions for administrationthereof preferably comprise instructions for administration thereof to apatient or subject in need thereof for improving the appearance of theskin, for example the periorbital skin. The formulation for improvingthe appearance of the skin comprising caffeine may also comprisetocopherol or a derivative thereof.

Where the AOI comprises tocopherol in the continuous phase and salicylicacid or a salt or ester thereof tethered to the deformable colloidalparticles, the instructions for administration thereof preferablycomprise instructions for administration thereof to a patient or subjectin need thereof for improving the appearance of the skin, for exampletreating and/or preventing uneven skin tone.

An eighth aspect of the present invention comprises a transdermal drugrelease device comprising a support layer and a layer comprising thecomposition according to the first aspect of the present invention.

The transdermal drug release device may comprise a strip, plaster,bandage or patch.

The support layer may be made of any suitable material including fabricand silicon.

The transdermal drug release device may be applied to the skin of apatient such that the layer of the composition of the first aspect ofthe invention is in contact with the skin of the patient.

The support layer may remain on the skin for a set period of time, forexample 1 hour to 3 days. Alternatively, the support layer may beremoved and the composition layer may remain in contact with the skin.

All preferred features of the first aspect of the invention also applyto the second, third, fourth, fifth, sixth, seventh and eighth aspectsof the invention.

Generally, the nomenclature used herein and the laboratory procedures inorganic chemistry, medicinal chemistry, and pharmacology describedherein are those well-known and commonly employed in the art. Unlessdefined otherwise, all technical and scientific terms used hereingenerally have the same meaning as commonly understood by one ofordinary skill in the art to which this disclosure belongs.

As used herein, “skin” includes the general skin of the body and anyother external integument, such as the epithelium of the ear, nose,throat and eye, including the sclera of the eye, and other mucosalmembranes, such as the vagina and anus/rectum.

As used herein, a “sufficient amount”, “amount effective to”, “amountsufficient to” or “amount of the formulation sufficient to” achieve aparticular result refers to an amount of the formulation of theinvention is effective to produce a desired effect, which is optionallya therapeutic effect (i.e., by administration of a therapeuticallyeffective amount). Alternatively stated, a “therapeutically effective”amount is an amount that provides some alleviation, mitigation, and/ordecrease in at least one clinical symptom. Clinical symptoms associatedwith the disorder that can be treated by the methods of the inventionare well-known to those skilled in the art. Further, those skilled inthe art will appreciate that the therapeutic effects need not becomplete or curative, as long as some benefit is provided to thesubject. For example, a “sufficient amount” or “an amount sufficient to”can be an amount that is effective to treat the symptoms of any diseaseor condition as herein described.

As used herein, the terms “treat”, “treating” or “treatment” of meanthat the severity of a subject's condition is reduced or at leastpartially improved or ameliorated and/or that some alleviation,mitigation or decrease in at least one clinical symptom is achievedand/or there is an inhibition or delay in the progression of thecondition and/or delay in the progression of the onset of disease orillness. The terms “treat”, “treating” or “treatment of” also meansmanaging the disease state.

As used herein, the term “pharmaceutically acceptable” when used inreference to the formulation of the invention denotes that a formulationdoes not result in an unacceptable level of irritation in the subject towhom the formulation is administered. Preferably such level will besufficiently low to provide a formulation suitable for approval byregulatory authorities.

As used herein with respect to numerical values, the term “about” or“approximately” means a range surrounding a particular numeral valuewhich includes that which would be expected to result from normalexperimental error in making a measurement. For example, in certainembodiments, the term “about” when used in connection with a particularnumerical value means +-20%, unless specifically stated to be +−1%,+−2%, +−3%, +−4%, +−5%, +−10%. +−15%, or +−20% of the numerical value.

In accordance with this disclosure, the term “comprising” is inclusiveor open-ended and does not exclude additional, un-recited elements ormethod steps; the term “consisting of” excludes any element, step, oringredient not specified; and the term “consisting essentially of”excludes any element, step, or ingredient that materially changes abasic characteristic of the invention.

The colloidal particles of the colloidal dispersions of the formulationof the invention may comprise a mixture of more than one non-ionicsurfactant. The colloidal particles of the colloidal dispersions of theformulation of the invention may comprise a mixture of more than onephospholipid. The colloidal particles of the colloidal dispersions ofthe formulation of the invention may comprise a mixture of more than onenon-ionic surfactant and more than one phospholipid. The colloidalparticles of the colloidal dispersions of the formulation of theinvention may comprise at least one non-ionic surfactant, at least onephospholipid, a pharmaceutically acceptable carrier, and optionallybuffers, antioxidants, preservatives, microbicides, antimicrobials,emollients, co-solvents, and/or thickeners. The colloidal particles ofthe colloidal dispersions of the formulation of the invention mayconsist essentially of a non-ionic surfactant and a phospholipid, apharmaceutically acceptable carrier, and optionally buffers,antioxidants, preservatives, microbicides, antimicrobials, emollients,co-solvents, and/or thickeners. The colloidal particles of the colloidaldispersions of the formulation of the invention may consist of at leastone non-ionic surfactant, at least one phospholipid, a pharmaceuticallyacceptable carrier, and one or more of the following: buffers,antioxidants, preservatives, microbicides, antimicrobials, emollients,co-solvents, and thickeners.

Any lipid known in the art that will form a phospholipid bilayer may beused for making the formulation of the invention.

Table 1 lists preferred phospholipids in accordance with the invention.

TABLE 1 Laur(o)yl Palmit(o)yl Stear(o)yl Bechen(o)yl Eruca(o)ylArachin(o)yl Gadolen(o)yl Arachindon(o)yl Ole(o)yl Linol(o)ylLinole(n/o)yl Palmitole(o)yl Myrist(o)yl Capr(o)yl

The preferred lipids in the context of this disclosure are uncharged andform stable, well hydrated bilayers; phosphatidylcholines,phosphatidylethanolamine, and sphingomyelins are the most prominentrepresentatives of such lipids. Any of those can have chains as listedin Table 1; the ones forming fluid phase bilayers, in which phospholipidchains are in disordered state, being preferred.

A preferred phospholipid is, for example, a natural phosphatidylcholine,which was formerly known as lecithin. It can be obtained from egg (richin palmitic, C16:0, and oleic, C18:1, but also comprising stearic,C18:0, palmitoleic, C16:1, linolenic, C18:2, and arachidonic, C20:4(M,radicals), soybean (rich in unsaturated C18 chains, but also containingsome palmitic radicals, amongst a few others), coconut (rich insaturated chains), olives (rich in monounsaturated chains), saffron(safflower) and sunflowers (rich in n-6 linoleic acid), linseed (rich inn-3 linolenic acid), from whale fat (rich in monounsaturated n-3chains), from primrose or primula (rich in n-3 chains). Preferred,natural phosphatidyl ethanolamines (used to be called cephalins)frequently originate from egg or soybeans. Preferred sphingomyelins ofbiological origin are typically prepared from eggs or brain tissue.Preferred phosphatidylserines also typically originate from brainmaterial whereas phosphatidylglycerol is preferentially extracted frombacteria, such as E. coli, or else prepared by way oftransphosphatidylation, using phospholipase D, starting with a naturalphosphatidylcholine. The preferably used phosphatidylinositols areisolated from commercial soybean phospholipids or bovine liver extracts.The preferred phosphatidic acid is either extracted from any of thementioned sources or prepared using phospholipase D from a suitablephosphatidylcholine.

Synthetic phosphatidylcholines, synthetic phosphatidylethanolamines,synthetic phosphatidic acids, synthetic phosphatidylserines, syntheticphosphatidyl(poly)alcohols, such as phosphatidylinositol, or syntheticphosphatidylglycerol may also be used in accordance with the invention.

Suitable lipids furthermore are a lysophosphatidyl choline analog, suchas 1-lauroyl-1,3-dihydroxypropane-3-phosphoryl choline, a monoglyceride,such as monoolein or monomyristin, a cerebroside, ceramide polyhexoside,sulfatide, sphingoplasmalogen, a ganglioside or a glyceride, which doesnot contain a free or esterified phosphoryl or phosphono or phosphinogroup in the 3 position. An example of such a glyceride isdiacylglyceride or 1-alkenyl-1-hydroxy-2-acyl glyceride with any acyl oralkenyl groups, wherein the 3-hydroxy group is etherified by one of thecarbohydrate groups named, for example, by a galactosyl group such as amonogalactosyl glycerin.

Lipids with desirable head or chain group properties can also be formedby biochemical means, for example, by means of phospholipases (such asphospholipase A1, A2, B, C and, in particular, D), desaturases,elongases, acyl transferases, etc., from natural or syntheticprecursors.

Furthermore, a suitable phospholipid is any phospholipid, which iscontained in biological membranes and can be extracted with the help ofapolar organic solvents, such as chloroform. Aside from the lipidsalready mentioned, such lipids also include, for example, steroids, suchas estradiol, or sterols, such as cholesterol, beta-sitosterol,desmosterol, 7-keto-cholesterol or beta-cholestanol, fat-solublevitamins, such as retinoids, vitamins, such as vitamin A1 or A2, vitaminE, vitamin K, such as vitamin K1 or K2 or vitamin D1 or D3, etc.

The less soluble amphiphilic components comprise or preferably comprisea synthetic phospholipid, such as myristoleoyl, palmitoleoyl,petroselinyl, petroselaidyl, oleoyl, elaidyl, cis- or trans-vaccenoyl,linolyl, linolenyl, linolaidyl, octadecatetraenoyl, gondoyl,eicosaenoyl, eicosadienoyl. eicosatrienoyl, arachidoyl, cis- ortrans-docosaenoyl, docosadienoyl, docosatrienoyl, docosatetraenoyl,lauroyl, tridccanoyl. myristoyl, pentadccanoyl, palmitoyl,heptadecanoyl, stearoyl or nonadecanoyl, glycerophospholipid orcorresponding derivatives with branched chains or a correspondingdialkyl or sphingosin derivative, glycolipid or other diacyl or dialkylphospholipid.

The more soluble amphiphilic components(s) is/are frequently derivedfrom the less soluble components listed above and, to increase thesolubility, substituted and/or complexed and/or associated with abutanoyl, pentanoyl, hexanoyl, heptanoyl, octanoyl, nonanoyl, decanoylor undecanoyl substituent or several, mutually independent, selectedsubstituents or with a different material for improving the solubility.

A further suitable phospholipid is a diacyl- ordialkyl-glycerophosphoethanolamine azo polyethoxylene derivative, adidecanoylphosphatidyl choline or a diacylphosphoolligomaltobionamide.

The amount of phospholipid in the formulations of the invention mayrange from about 0% to about 12%, about 0.2% to about 10%, about 0.6% toabout 8%, about 1.0% to about 7%, about 1.5% to about 6%, about 2% toabout 4%, about 3% to about 3.5% or about 0.7%, about 1.4%, about 3.4%or about 3.6% by weight. Preferably, the phospholipid is aphosphatidylcholine.

The formulations of the invention may include a surfactant. The term“surfactant” has its usual meaning. A list of relevant surfactants andsurfactant related definitions is provided in EP 0 475 160 A1 (see,e.g., p. 6, 1. 5 to p.14. 1.17) and U.S. Pat. No. 6,165,500 (see, e g.,col. 7, 1. 60 to col. 19, 1. 64), and in appropriate surfactant orpharmaceutical Handbooks, such as Handbook of Industrial Surfactants orUS Pharmacopoeia, Pharm. Eu. The surfactants may be those described inTables 1-18 of U.S. Patent Application Publication No. 2002/0012680 A1,published Jan. 31, 2002. The following list therefore only offers aselection, which is by no means complete or exclusive, of severalsurfactant classes that are particularly common or useful in conjunctionwith present patent application. Preferred surfactants to be used inaccordance with the disclosure include those with an HLB greater than12. The list includes ionized long-chain fatty acids or long chain fattyalcohols, long chain fatty ammonium salts, such as alkyl- oralkenoyl-trimethyl-, -dimethyl- and -methyl-ammonium salts, alkyl- oralkenoyl-sulphate salts, long fatty chain dimethyl-aminoxides, such asalkyl- or alkenoyl-dimethyl-aminoxides, long fatty chain, for examplealkanoyl, dimethyl-aminoxides and especially dodecyl dimethyl-aminoxide,long fatty chain, for example alkyl-N-methylglucamide-s andalkanoyl-N-methylglucamides, such as MEGA-8, MEGA-9 and MEGA-IO, N-longfatty chain-N,N-dimethylglycines, for exampleN-alkyl-N,N-dimethylglycines, 3-(long fattychain-dimethylammonio)-alkane-sulphonates, for example3-(acyidimethylammonio)-alkanesulphonates, long fatty chain derivativesof sulphosuccinate salts, such as bis(2-ethylalkyl)sulphosuccinatesalts, long fatty chain-sulphobetaines, for example acyl-sulphobetaines,long fatty chain betaines, such as EMPIGEN BB or ZWITTERGENT-3-16,-3-14, -3-12, -3-10, or -3-8, or polyethylcn-glycol-acylphenyl ethers,especially nonaethylen-glycol-octyl-phenyl ether, polyethylene-longfatty chain-ethers, especially polyethylene-acyl ethers, such asnonaethylen-decyl ether, nonaethylen-dodecyl ether oroctaethylene-dodecyl ether, polyethyleneglycol-isoacyl ethers, such asoctaethyleneglycol-isotridecyl ether, polyethyleneglycol-sorbitane-longfatty chain esters, for example polyethyleneglycol-sorbitane-acyl estersand especially polyoxyethylene-monolaurate (e.g. polysorbate 20 or Tween20), polyoxyethylene-sorbitan-monooleate (e.g.

polysorbate 80 or Tween 80), polyoxyethylene-sorbitan-monolauroleylate,polyoxyethylene—sorbitan-monopetroselinate,polyoxyethylene-sorbitan-monoelaidate,polyoxyethylene—sorbitan-myristoleylate, polyoxyethylene-sorbitan-palmitoleinylate, polyoxyethylene-sorbitan-p-etroselinylate,polyhydroxyethylene-long fatty chain ethers, for examplepolyhydroxyethylene-acyl ethers, such as polyhydroxyethylene-laurylethers, polyhydroxyethylene-myristoyl ethers,polyhydroxyethylene-cetylst-earyl, polyhydroxyethylene-palmityl ethers,polyhydroxyethylene-oleoyl ethers, polyhydroxyethylene-palmitoleoylethers, polyhydroxyethylene-lino-leyl, polyhydroxyethylen-4, or 6, or 8,or 10, or 12-lauryl, miristoyl, palmitoyl, palmitoleyl, oleoyl orlinoeyl ethers (Brij series), or in the corresponding esters,polyhydroxyethylen-laurate, -myristate, -palmitate, -stearate or-oleate, especially polyhydroxyethylen-8-stearate (Myrj 45) andpolyhydroxyethylen-8-oleate, polyethoxylated castor oil 40 (CremophorEL), sorbitane-mono long fatty chain, for example alkylate (Arlacel orSpan series), especially as sorbitane-monolaurate (Arlacel 20, Span 20),long fatty chain, for example acyl-N-methylglucamides,alkanoyl-N-methylglucamides, especially decanoyl-N-methylglucamide,dodecanoyl-N-methylglucamide, long fatty chain sulphates, for examplealkyl-sulphates, alkyl sulphate salts, such as lauryl-sulphate (SDS),oleoyl-sulphate: long fatty chain thioglucosides, such asalkylthioglucosides and especially heptyl-, octyl- andnonyl-beta-D-thioglucopyranoside; long fatty chain derivatives ofvarious carbohydrates, such as pentoses, hcxoses and disaccharidcs,especially alkyl-glucosides and maltosides, such as hexyl-, heptyl-,octyl-, nonyl- and decyl-beta-D-glucopyranoside or D-maltopyranosidc;further a salt, such as a fatty acid salt, especially oleate, elaidate,linoleate, laurate, or myristate, most often in sodium form,lysophospholipids, n-octadecylene-glycerophosphatidic acid,octadecylene-phosphorylglycerol, octadecylene-phosphorylserine, n-longfatty chain-glycero-phosphatidic acids, such asn-acyl-glycero-phosphatidic acids, especially laurylglycero-phosphatidic acids, oleoyl-glycero-phosphatidic acid, n-longfatty chain-phosphoryl glycerol, such as n-acyl-phosphorylglycerol,especially lauryl-, myristoyl-, oleoyl- orpalmitoeloyl-phosphorylglycerol, n-long fatty chain-phosphorylserine,such as n-acyl-phosphorylserine, especially lauryl-, myristoyl-, oleoyl-or palmitoeloyl-phosphorylserine, n-tetradecyl-glycero-phosphatidicacid, n-tetradecyl-phosphorylglycerol, n-tetradecyl-phosphorylserine,corresponding-, elaidoyl-, vaccenyl-lysophospholipids, correspondingshort-chain phospholipids, as well as all surface active and thusmembrane destabilising polypeptides. Surfactant chains are typicallychosen to be in a fluid state or at least to be compatible with themaintenance of fluid-chain state in carrier aggregates.

The surfactant in the formulations of the invention is a non-ionicsurfactant. The surfactant may be present in the formulation in about 0%to about 1%, about 0.04% to about 0.8%, about 0.06% to about 0.6%, about0.08% to about 0.4%, about 0.1% to about 0.3%, about 0.15% to about0.2%, about 0.1% to about 5%, about 0.2% to about 3%, about 0.3% toabout 4%, about 0.5% to about 10%, about 1% to about 9%, about 1.5% toabout 7%, about 2% to about 6% or about 3% to about 8% by weight. Thenon-ionic surfactant may be selected from the group consisting of:polyoxyethylene sorbitans (polysobate surfactants), polyhydroxyethylenestearates or polyhydroxyethylene laurylethers (Brij surfactants). Thesurfactant may be a polyoxyethylene-sorbitan-monooleate (e.g.polysorbate 80 or Tween 80) or Tween 20, 40 or 60. The polysorbate canhave any chain with 12 to 20 carbon atoms. The polysorbate may be fluidin the formulations, which may contain one or more double bonds,branching, or cyclo-groups.

The formulations of the invention may comprise only one non-ionicsurfactant. The formulations of the invention may comprise only onephospholipid. The formulations of the invention may comprise only onenon-ionic surfactant and only one phospholipid. The formulations of theinvention may comprise one or more than one non-ionic surfactant and/orone or more than one phospholipid, e.g., independently one, two, three,four, or more non-ionic surfactants and/or one, two, three, four or morephospholipids.

The formulations of the invention may have a range of lipid/phospholipidto surfactant ratios. The ratios may be expressed in terms of molarterms (mol lipid/mol surfactant or mol phospholipid/mol surfactant).They may also be expressed as % weight. The weight ratio of lipid orphospholipid to surfactant in the formulation may be from about 20:1 toabout 5:1, from about 15:1 to about 10:1, from about 12:1 to about 9:1,from about 18:1 to about 14:1. Specifically, the weight ratio of lipidor phospholipid to surfactant in the formulation may be about 8:1, about9:1, about10:1, about 15:1 or about 16:1. There may be no lipid orphospholipid present i.e. a ratio of 0:1. There may be no surfactantpresent i.e. a ratio of 1:0. Preferably, all of the deformable colloidalparticles in a formulation of the present invention have the same lipidor phospholipid to surfactant ratio.

The formulations of the invention may also have varying amounts of totalamount of the following components: lipid or phospholipid and surfactantcombined (TA). The TA amount may be stated in terms of weight percent ofthe total composition. The TA may be from about 1% to about 40%, about5% to about 30%, about 7.5% to about 15%, about 6% to about 14%, about8% to about 12%, about 5% to about 10%, about 10% to about 20% or about20% to about 30%. The TA may be 6%, 7%, 8%, 9%, 10%, 12%, 14%, 15% or20%.

The molar ratio of lipid or phospholipid to surfactant in theformulations of the invention may be from about 1:3 to about 30:0, fromabout 1:3 to about 30:1, from about 1:2 to about 30:1, from about 1:1 toabout 30:0, from about 1:1 to about 30:1, from about 2:1 to about 20:0,from about 2:1 to about 20:1, from about 5:1 to about 30:1, from about10:1 to about 30:1, from about 15:1 to about 30:1, or from about 20:1 toabout 30:1. The molar ratio of lipid or phospholipid to surfactant inthe formulation may be from about 1:2 to about 10:1. The molar ratio maybe from about 1:1 to about 2:1, from about 2:1 to about 3:1, from about3:1 to about 4:1, from about 4:1 to about 5:1 or from about 5:1 to about10:1. The molar ratio may be from about 10.1 to about 30:1, from about10:1 to about 20:1, from about 10:1 to about 25:1, and from about 20:1to about 25:1. The lipid or phospholipid to surfactant ratio may beabout 1.0:1.0, about 1.25:1.0, about 1.5:1.0, about 1.75:1.0, about2.0:1.0, about 2.5:1.0, about 3.0:1.0 or about 4.0:1.0.

Selected ranges for total lipid or phospholipid amounts and lipid orphospholipid/surfactant ratios (mol/mol) for the formulations of theinvention are described in Table 2 below:

TABLE 2 Total Amount and Lipid/Phospholipid to Surfactant Ratios.Lipid/Surfactant or Phospholipid/Surfactant TA (%) (mol/mol) 5 to 10 1.0to 1.25 5 to 10 1.25 to 1.72 5 to 10 1.75 to 2.25 5 to 10 2.25 to 3.00 5to 10 3.00 to 4.00 5 to 10 4.00 to 8.00 5 to 10 10.00 to 13.00 5 to 1015.00 to 20.00 5 to 10 20.00 to 22.00 5 to 10 22.00 to 25.00 10 to 201.0 to 1.25 10 to 20 1.25 to 1.75 10 to 20 1.25 to 1.75 10 to 20 2.25 to3.00 10 to 20 3.00 to 4.00 10 to 20 4.00 to 8.00 10 to 20 10.00 to 13.0010 to 20 15.00 to 20.00 10 to 20 20.00 to 22.00 10 to 20 22.00 to 25.00

The formulations of the invention may optionally contain one or more ofthe following ingredients: co-solvents, chelators, buffers,antioxidants, preservatives, microbicides, emollients, humectants,lubricants and thickeners. Preferred amounts of optional components aredescribed as follows in Table 3.

TABLE 3 Molar (M) or Rel w %* Antioxidant: Primary: Butylatedhydroxyanisole, BHA 0.1-8 Butylated hydroxytoluene BHT 0.1-4 Thymol0.1-1 Metabisulphite 1-5 mM Bisulsphite 1-5 mM Thiourea (MW = 76.12)1-10 mM Monothioglycerol (MW = 108.16) 1-20 mM Propyl gallate (MW =212.2)  0.02-0.2 Ascorbate (MW = 175.3⁺ ion) 1-10 mM Palmityl-ascorbate0.01-1  Tocopherol-PEG 0.5-5 Secondary (chelator) EDTA (MW = 292) 1-10mM EGTA (MW = 380.35) 1-10 mM Desferal (MW = 656.79) 0.1-5 mM BufferAcetate 30-150 mM Phosphate 10-50 mM Triethanolamine 30-150 mM *as apercentage of total phospholipid quantity

The formulation of the invention is typically formulated and/oradministered in an aqueous medium. The formulation may be formulatedwith or without co-solvents, such as lower alcohols. The formulation ofthe present invention may also comprise a polar liquid medium. Theformulation may be in the form of a solution, suspension, emulsion,cream, lotion, ointment, gel, spray, film forming solution or lacquer.Preferably, the formulation of the present invention is in the form of agel.

The formulation of the invention may include a buffer to adjust the pHof the aqueous solution to a range from pH 3.5 to pH 9, pH 4 to pH 7.5,or pH 6 to pH 7. Examples of buffers include, but are not limited toacetate buffers, lactate buffers, phosphate buffers, and propionatebuffers. Preferably, the formulations comprise one or more buffersselected from the group consisting of disodium hydrogen orthophosphatedodecahydrate, disodium hydrogen orthophosphate anhydrous, sodiumdihydrogen orthophosphate dehydrate, sodium dihydrogen orthophosphatedodecahydrate and phosphate buffer, for example phosphate (pH6.7)buffer.

A “microbicide” or “antimicrobial”' agent is commonly added to reducethe bacterial count in pharmaceutical formulations. Some examples ofmicrobicides are short chain alcohols, including ethyl and isopropylalcohol, chlorbutanol, benzyl alcohol, chlorbenzyl alcohol,dichlorbenzylalcohol, hexachlorophene; phenolic compounds, such ascresol, 4-chloro-m-cresol, p-chloro-m-xylenol, dichlorophene,hexachlorophene, povidon-iodine; parabenes, especially alkyl-parabenes,such as methyl-, ethyl-, propyl-, or butyl-paraben, benzyl paraben;acids, such as sorbic acid, benzoic acid and their salts; quaternaryammonium compounds, such as alkonium salts, e.g., a bromide,benzalkonium salts, such as a chloride or a bromide, cetrimonium salts,e.g., a bromide, phenoalkecinium salts, such as phenododecinium bromide,cetylpyridinium chloride and other salts; furthermore, mercurialcompounds, such as phenylmercuric acetate, borate, or nitrate,thiomersal, chlorhexidine or its gluconate, or any antibiotically activecompounds of biological origin, or any suitable mixture thereof.

Examples of “antioxidants” are butylhydroxyanisole or butylatedhydroxyanisol (BHA), butylated hydroxytoluene (BHT) anddi-tert-butylphenol (LY178002, LY256548, HWA-131, BF-389, Cl-986,PD-127443, E-51 or 19, Bl-L-239XX, etc.), tertiary butylhydroquinone(TBHQ), propyl gallate (PG), I-O-hexyl-2,3,5-trimethylhydroquinone(HTHQ); aromatic amines (diphenylamine, p-alkylthio-o-anisidine,ethylenediamine derivatives, carbazol, tetrahydroindenoindol); phenolsand phenolic acids (guaiacol, hydroquinone, vanillin, gallic acids andtheir esters, protocatechuic acid, quinic acid, syringic acid, ellagicacid, salicylic acid, nordihydroguaiaretic acid (NDGA), eugenol);tocopherols (including tocopherols (alpha, beta, gamma, delta) and theirderivatives, such as tocopheryl-acylate (e g. -acetate, -laurate,myristate, -palmitate, -oleate, -linoleate, etc., or an y other suitabletocopheryl-lipoate), tocopheryl-POE-succinate; trolox and correspondingamide and thiocarboxamide analogues; ascorbic acid and its salts,isoascorbate, (2 or 3 or 6)-o-alkylascorbic acids, ascorbyl esters(e.g., 6-o-lauroyl, myristoyl, palmitoyl-, oleoyl, orlinoleoyl-L-ascorbic acid, etc.). Also useful are the preferentiallyoxidised compounds, such as sodium bisulphite, sodium metabisulphite,thiourea; chellating agents, such as EDTA, GDTA, desferral:miscellaneous endogenous defence systems, such as transferrin,lactoferrin, ferritin, cearuloplasmin, haptoglobion, heamopexin,albumin, glucose, ubiquinol-10); enzymatic antioxidants, such assuperoxide dismutase and metal complexes with a similar activity,including catalase, glutathione peroxidase, and less complex molecules,such as beta-carotene, bilirubin, uric acid; flavonoids (flavones,flavonols, flavonones, flavanonals, chacones, anthocyanins).N-acetylcystein, mesna. glutathione, thiohistidine derivatives,triazoles; tannines, cinnamic acid, hydroxycinnamatic acids and theiresters (coumaric acids and esters, caffeic acid and their esters,ferulic acid, (iso-) chlorogenic acid, sinapic acid);

spice extracts (e.g., from clove, cinnamon, sage, rosemary, mace,oregano, allspice, nutmeg); carnosic acid, carnosol, carsolic acid;rosmarinic acid, rosmaridiphenol, gentisic acid, ferulic acid; oat flourextracts, such as avenanthramide 1 and 2; thioethers, dithioethers,sulphoxides, tetralkylthiuram disulphides; phytic acid, steroidderivatives (e.g., U74006F); tryptophan metabolites (e.g.,3-hydroxykynurenine, 3-hydroxyanthranilic acid), andorganochalcogenides.

“Thickeners” are used to increase the viscosity of pharmaceuticalformulations and may be selected from pharmaceutically acceptablehydrophilic polymers, such as partially etherified cellulosederivatives, comprising carboxym ethyl-, hydroxyethyl-, hydroxypropyl-,hydroxypropylmethyl- or methyl-cellulose; completely synthetichydrophilic polymers such as polyacrylates (e.g. Carbopol®, specificallycarbopol 974P), polymethacrylatcs, poly(hydroxyethyl)-,poly(hydroxypropyl)-, poly(hydroxypropylmethyl)methacrylate,polyacrylonitrile, methallyl-sulphonate, polyethylenes,polyoxiethylenes, polyethylene glycols, polyethylene glycol-lactide,polyethylene glycol-diacrylate, polyvinylpyrrolidone, polyvinylalcohols, poly(propylmethacrylamide), poly(propylenefumarate-co-ethylene glycol), poloxamers, polyaspartamide (hydrazinecross-linked) hyaluronic acid, silicone; natural gums comprisingalginates, carrageenan, guar-gum, gelatine, tragacanth, (amidated)pectin, xanthan, chitosan collagen, agarose; mixtures and furtherderivatives or co-polymers thereof and/or other pharmaceutically, or atleast biologically, acceptable polymers. Preferably, the thickeners usedin the formulations of the present invention may be selected fromCarbopol® (Carbopol® 974P NF), Methylcellulose (Metolose SM 4000),Keltrol CG-T (a personal care grade xanthan gum), Xantural 11k (apharmaceutical grade xanthan gum), Hydroxypropyl ethylcellulose,Genuvisco CG-131 (carrageenan gum), Kelcogel (gellan gum), SucraclearHC-31 (cellulose gum, carrageenan, Ceratonia siliqua gum, sucrose),Sucrathix VX (microcrystalline cellulose, cellulose gum, xanthan gum),Lubrajel (glyceryl acrylate/acrylic acid) or Chitosan.

One particularly preferred formulation of the present inventioncomprises insoluble aggregates or micelles of chlorhexidine or a saltthereof, preferably chlorhexidine digluconate, together with emptydeformable colloidal particles, preferably empty Sequessomes™, anddeformable colloidal particles, preferably Tethersomes, to which zincstearate has been tethered.

Another particularly preferred formulation of the present inventioncomprises liposomes comprising capsaicin, menthol and phospholipidstogether with deformable colloidal particles, preferably Transfersomes™,the membranes of which comprise menthol.

A particularly preferred formulation comprises myristyl salicylate ortridecyl salicylate dissolved in the continuous phase, and deformablecolloidal particles, preferably Tethersomes, to which myristylsalicylate or tridecyl salicylate are tethered. Preferably, a derivativeof tocopherol is also tethered to said deformable colloidal particles.This composition additionally comprises a second type of deformablecolloidal particle, preferably Tethersomes, to which palmitoyl ascorbicacid and tocopheryl linoleate are tethered. Tocopherol may be present inthe continuous phase of the formulation.

Another particularly preferred formulation comprises glucosaminehydrochloride and chondroitin sulphate dissolved in the continuousphase, and deformable colloidal particles, preferably Tethersomes, towhich palmitoyl ascorbate and tocopheryl linoleate are preferablytethered.

A further particularly preferred formulation comprises N-acetylglucosamine sulphate and chondroitin sulphate dissolved in thecontinuous phase, and deformable colloidal particles, preferablyTethersomes, to which palmitoyl ascorbic acid and tocopheryl linoleateare preferably tethered.

Another particularly preferred composition comprises caffeine dissolvedin the continuous phase, and deformable colloidal particles, preferablyTethersomes, to which palmitoyl ascorbic acid and tocopheryl linoleateare preferably tethered. This composition additionally comprises twofurther types of deformable colloidal particles, a first to whichpalmitoyl tripeptide-1 is tethered and a second to which palmitoyltetrapeptide-7 is tethered. Preferably, the composition comprisestocopherol as a separate phase. A small amount of tocopherol maypartition into the lumen of the deformable colloidal particles. Oneembodiment of such a formulation is set out in Example 6.

A further particularly preferred formulation comprises deformablecolloidal particles, which comprise tridecyl salicylate and/or alphatocopherol. Tridecyl salicylate is preferably tethered to the colloidalparticles. Alpha tocopherol may be present in the continuous phase ofthe formulations (i.e. not associated with the colloidal particles) andsome may partition into the lumen of the colloidal particles. Thiscomposition additionally comprises a further type of deformablecolloidal particle, which comprises palmitoyl ascorbic acid, tocopheryllinoleate and/or alpha tocopherol. Palmitoyl ascorbic acid andtocopheryl linoleate are preferably tethered to the colloidal particle.Alpha tocopherol may be present in the continuous phase of theformulations (i.e. not associated with the colloidal particles) and somemay partition into the lumen of the colloidal particles. One embodimentof such a formulation is set out in Example 7.

BRIEF DESCRIPTION OF THE DRAWINGS

The invention is described below with reference to the followingexamples and figures in which:

FIG. 1 shows the structure of a colloidal particle (vesicle orSequessome™/Transfersome™/Tethersome) of the invention. The sphericalbilayer may comprise a surfactant or a lipid or a surfactant and alipid. As can be seen from this diagram, the bilayer is two moleculeswide;

FIG. 2 shows a typical formulation in accordance with the first aspectof the invention. This formulation comprises a colloidal dispersion (orSequessome™/Transfersome™/Tethersome intermediate) comprising colloidalparticles, a colloidal dispersion comprising colloidal particlescomprising an agent of interest (AOI) (e.g. a peptide, ascorbic acid orzinc) and an AOI that is not associated with the colloidal particles(e.g. chlorhexidine or capsaicin);

FIG. 3 shows a table summarising Example Formulations 1 to 4;

FIG. 4 shows the change in sebum levels compared to baseline level onday 0 (=100%);

FIG. 5 shows the change in percentage of comedones compared to day 0occurrence (100%);

FIG. 6 shows the change in percentage incidence of pustules compared today 0 occurrence (100%);

FIG. 7 shows the reduction in measured wrinkle volume and the reductionin appearance of fine lines and wrinkles after treatment with a twotypes of formulation in accordance with the present invention;

FIG. 8 shows a comparison of reduction in wrinkle volume (as apercentage of Week 0 baseline volume) after treatment with a formulationin accordance with the present invention, a UK leading own brand(in-market product) and a control;

FIG. 9 shows a comparison of reduction in perceived appearance ofwrinkles and fine lines (as a percentage of Week 0 baseline volume)after treatment with a formulation in accordance with the presentinvention, a UK leading own brand (in-market product) and a control; and

FIG. 10 shows a comparison of reduction in appearance of pen-orbitaldark circles (as a percentage of Week 0 baseline volume) after treatmentwith a formulation in accordance with the present invention, a UKleading own brand (in-market product) and a control.

EXAMPLES Example 1: Method of Manufacture

The simplified generic manufacturing process of formulations inaccordance with the present invention is as follows. Firstly, thecolloidal dispersions comprising the deformable colloidal particles (forexample, the Sequessome™ intermediate, Transfersome™ intermediate orTethersome intermediate) are made. These colloidal dispersions are eachformed from an ‘organic phase’ containing alcohol-soluble components andan ‘aqueous phase’ consisting of water-soluble components. Secondly, thegel phase (a thickener) is formed within which the colloidaldispersion(s) are dispersed. During this secondary stage, more than onekind of colloidal dispersion is introduced (each kind containingdiffering AOIs (or ‘extra’ ingredients) associated with the colloidalparticles to give them unique characteristics), such that the finalgelled product is then a blend of one or more individual kinds ofvesicles (or colloidal particles). The AOIs or ‘extras’ may include zincstearate, palmitoyl peptides, palmitoyl ascorbic acid (PAA), myristylsalicylate, tridecyl salicylate, vitamin D or vitamin E etc. (the‘tethered’ ingredients). If desired, an AOI not associated with thecolloidal particles can be added to the gel phase during secondarymanufacture. If desired the first phase of the manufacturing process canbe ordered so that the colloidal dispersions comprising the colloidalparticles without any AOI are made in one or more formulations and thento each formulation an AOI (attached to a [phospho]lipid or asurfactant) is added. The second gel phase then occurs as above.

Example 2: Example Formulations Example Formulation 1

Quantity Required Per 100 g Final Product (g) Organic Phase - 1600 ppmPAA Intermediate SPC 4.12200 Ethanol 2.19660 BHT 0.01200Methyl-4-hydroxybenzoate 0.15000 Ethyl-4-hydroxybenzoate 0.15000 Benzylalcohol 0.31500 Polysorbate 80 0.40800 PAA 0.09600 Linalool 0.06000Organic Phase Sub-Total 7.50960 Aqueous Phase - 1600 ppm PAAIntermediate Sodium metabisulphite 0.03000 Citric acid 0.04320 Na EDTA0.06000 diNa hydrogen phos 12H2O 0.17280 Water 27.18420 Aqueous PhaseSub-Total 27.49020 PAA Tethersome Intermediate Total 34.99980 OrganicPhase - 300 ppm Tetrapeptide Intermediate SPC 1.37400 Ethanol 0.74120BHT 0.0040 Methyl-4-hydroxybenzoate 0.05000 Ethyl-4-hydroxybenzoate0.05000 Benzyl alcohol 0.10500 Polysorbate 80 0.15300 Palmitoyl peptide0.00600 Linalool 0.02000 Organic Phase Sub-Total 2.50320 Aqueous Phase -300 ppm Tetrapeptide Intermediate Sodium metabisulphite 0.01000 Citricacid 0.01440 Na EDTA 0.02000 diNa hydrogen phos 12H2O 0.05760 Water9.06140 Aqueous Phase Sub-Total 9.16340 Tetrapeptide TethersomeIntermediate Total 11.66660 Organic Phase - 300 ppm TripeptideIntermediate SPC 1.37400 Ethanol 0.74120 BHT 0.00400Methyl-4-hydroxybenzoate 0.05000 Ethyl-4-hydroxybenzoate 0.05000 Benzylalcohol 0.10500 Polysorbate 80 0.15300 Palmitoyl peptide 0.00600Linalool 0.02000 Organic Phase Sub-Total 2.50320 Aqueous Phase - 300 ppmTripeptide Intermediate Sodium metabisulphite 0.01000 Citric acid0.01440 Na EDTA 0.02000 diNa hydrogen phos 12H2O 0.05760 Water 9.06140Aqueous Phase Sub-Total 9.16340 Tripeptide Tethersome Intermediate Total11.66660 Sum of Tethersome Intermediates 58.33300 Gel Phase - FinalProduct Sodium hydroxide 0.11300 Carbopol 974P 0.75000 Glycerol 3.00000Citric acid 0.05600 diNa hydrogen phos 12H2O 0.24200 Water 37.50600 GelPhase Sub-Total 41.66700 Overall Total 100.00000

Example Formulation 2

Quantity Required Per 100 g Final Product (g) Organic Phase - 1600 ppmPAA Intermediate SPC 2.06100 Ethanol 1.09830 BHT 0.00600Methyl-4-hydroxybenzoate 0.07500 Ethyl-4-hydroxybenzoate 0.07500 Benzylalcohol 0.15750 Polysorbate 80 0.20400 PAA 0.04800 Linalool 0.03000Organic Phase Sub-Total 3.75480 Aqueous Phase - 1600 ppm PAAIntermediate Sodium metabisulphite 0.01500 Citric acid 0.02160 diNahydrogen phos 12H2O 0.08640 Water 13.62210 Aqueous Phase Sub-Total13.74510 PAA Tethersome Intermediate Total 17.49990 Organic Phase - 300ppm Tetrapeptide Intermediate SPC 0.68700 Ethanol 0.37060 BHT 0.00200Methyl-4-hydroxybenzoate 0.02500 Ethyl-4-hydroxybenzoate 0.02500 Benzylalcohol 0.05250 Polysorbate 80 0.07650 Palmitoyl peptide 0.00300Linalool 0.01000 Organic Phase Sub-Total 1.25160 Aqueous Phase - 300 ppmTetrapeptide Intermediate Sodium metabisulphite 0.00500 Citric acid0.00720 diNa hydrogen phos 12H2O 0.02880 Water 4.54070 Aqueous PhaseSub-Total 4.58170 Tetrapeptide Tethersome Intermediate Total 5.83330Organic Phase - 300 ppm Tripeptide Intermediate SPC 0.68700 Ethanol0.37060 BHT 0.00200 Methyl-4-hydroxybenzoate 0.02500Ethyl-4-hydroxybenzoate 0.02500 Benzyl alcohol 0.05250 Polysorbate 800.07650 Palmitoyl peptide 0.00300 Linalool 0.01000 Organic PhaseSub-Total 1.25160 Aqueous Phase - 300 ppm Tripeptide Intermediate Sodiummetabisulphite 0.00500 Citric acid 0.00720 diNa hydrogen phos 12H2O0.02880 Water 4.54070 Aqueous Phase Sub-Total 4.58170 TripeptideTethersome Intermediate Total 5.83330 Organic Phase - Zinc StearateIntermediate SPC 3.43500 Ethanol 1.82850 BHT 0.01000Methyl-4-hydroxybenzoate 0.12500 Ethyl-4-hydroxybenzoate 0.12500 Benzylalcohol 0.26250 Polysorbate 80 0.34000 Zinc Stearate 0.08200 Linalool0.05000 Organic Phase Sub-Total 6.25800 Aqueous Phase - Zinc StearateIntermediate Sodium metabisulphite 0.02500 Citric acid 0.03600 diNahydrogen phos 12H2O 0.14400 Water 22.70350 Aqueous Phase Sub-Total22.90850 Zinc Tethersome Intermediate Total 29.16650 Sum of TethersomeIntermediates 58.33300 Gel Phase - Final Product Sodium hydroxide0.11300 Carbopol 974P 0.75000 Glycerol 3.00000 Citric acid 0.05600 diNahydrogen phos 12H2O 0.24200 Water 37.50600 Gel Phase Sub-Total 41.66700Overall Total 100.00000

Example Formulation 3

Quantity Required Per 100 g Final Product (g) Organic Phase - EmptySequessome Intermediate SPC 3.57300 Ethanol 1.75000 BHA 0.01000Methyl-4-hydroxybenzoate 0.12500 Ethyl-4-hydroxybenzoate 0.12500 Benzylalcohol 0.26250 Polysorbate 80 0.23600 Linalool 0.05000 Organic PhaseSub-Total 6.13150 Aqueous Phase - Empty Sequessome Intermediate Nadihydrogen phos 2H2O 0.22200 diNa hydrogen phos 12H2O 0.38600 Water34.02100 Aqueous Phase Sub-Total 34.62900 Empty Sequessome IntermediateTotal 40.76050 Organic Phase - Zinc Stearate Intermediate SPC 3.57300Ethanol 1.66800 BHA 0.01000 Methyl-4-hydroxybenzoate 0.12500Ethyl-4-hydroxybenzoate 0.12500 Benzyl alcohol 0.26250 Polysorbate 800.23600 Zinc stearate 0.08200 Linalool 0.05000 Organic Phase Sub-Total6.13150 Aqueous Phase - Zinc Stearate Intermediate Na dihydrogen phos2H2O 0.22200 diNa hydrogen phos 12H2O 0.38600 Water 34.02100 AqueousPhase Sub-Total 34.62900 Zinc Tethersome Intermediate Total 40.76050 Sumof Sequessome and 81.52100 Tethersome lntermediates Gel Phase - FinalProduct Sodium hydroxide 0.30000 Carbopol 974P 1.00000 Glycerol 3.00000Water 14.17900 Gel Phase Sub-Total 18.47900 Overall Total 100.00000

Example Formulation 4

Quantity Required Per 100 g Final Product (g) Organic Phase - EmptySequessome Intermediate Soy phosphatidylcholine (SPC) 3.43500 Ethanol1.82550 BHA 0.01000 Methyl-4-hydroxybenzoate 0.12500Ethyl-4-hydroxybenzoate 0.12500 Benzyl alcohol 0.26250 Polysorbate 800.42500 Linalool 0.05000 Organic Phase Sub-Total 6.25800 Aqueous Phase -Empty Sequessome Intermediate Citric acid monohydrate 0.03600 diNahydrogen phos 12H2O 0.14400 Water 22.72850 Aqueous Phase Sub-Total22.90850 Empty Sequessome Intermediate Total 29.16650 Organic Phase -Zinc Stearate Intermediate Soy phosphatidylcholine SPC 3.43500 Ethanol1.74350 BHA 0.01000 Methyl-4-hydroxybenzoate 0.12500Ethyl-4-hydroxybenzoate 0.12500 Benzyl alcohol 0.26250 Polysorbate 800.42500 Zinc stearate 0.08200 Linalool 0.05000 Organic Phase Sub-Total6.25800 Aqueous Phase - Zinc Stearate Intermediate Citric acidmonohydrate 0.03600 diNa hydrogen phos 12H2O 0.14400 Water 22.72850Aqueous Phase Sub-Total 22.90850 Zinc Tethersome Intermediate Total29.16650 Sum of Sequessome and 58.33300 Tethersome Intermediates GelPhase - Final Product Citric acid monohydrate 0.05600 diNa hydrogen phos12H2O 0.24200 Methyl cellulose (4000 cPs, 88,000 Mw) 1.75000 Glycerol3.00000 Water 36.00600 Gel Phase Sub-Total 40.16700 Chlorhexidinedigluconate 1.50000 Overall Total 100.00000

Example 3

An in-use study in healthy volunteers to investigate the anti-spotefficacy of two Example Formulations comprising chlorhexidine, usingobjective instrumental assessments of skin sebum levels and subjectivevisual assessments of lesion prevalence, against a placebo following a3-week use period.

The effectiveness of Formulation 4 and a second formulation(“Formulation 4.2”) was compared with a control product in their abilityto control sebum and reduce the incidence of acne in acne-proneindividuals.

The Example Formulations (n=36 and n=37) contained empty Sequessomes™and Tethersomes with tethered zinc and other excipients, includingchlorhexidine as an anti-microbial preservative. The Sequessomes™ in thetwo Formulations contained different ratios of SPC:Tween (polysorbate)to assess if this affected the efficacy of the product. The controlproduct contained no vesicles, zinc or chlorhexidine (n=20).

Data Review

The study assessed the efficacy and safety of treatment with Formulation4 and Formulation 4.2 compared with placebo (control) in healthyvolunteers with oily and spot-prone skin, confirmed by a clinical trialsassistant prior to enrolment.

Volunteers were either assessed at the centre on Day 3, 7, 14 and 21 orpurely provided their own subjective assessments from home.

In total, 224 volunteers completed the 3-week study: 37 were allocatedto Formulation 4, 36 were allocated to Formulation 4.2 and 20 toplacebo. Objective assessments included measurement of sebum on theface, Visia CR Photography (used to assess pore size and skinsmoothness) and lesion counts (comedones, microcysts, papules andpustules). The subjective assessment was carried out using a subjectSelf Perception Questionnaire (SPQ).

Sebum levels were reduced by over 20% from Baseline after 1 week ofFormulation 4 and by more than 50% after 3 weeks' treatment (see FIG.4). Published data demonstrate that a reduction in sebum production of30-50% correlates with reduced acne symptoms.¹ The number of comedones(blocked pores) was also reduced with Formulation 4: by 50% fromBaseline after 1 week and by more than 80% after 3 weeks (see FIG. 5).Notably, the number of pustules was reduced by 90% from Baseline after 1week and they were virtually cleared after 3 weeks' treatment (see FIG.6).

These objective measurements were further validated by the subjectiveassessments. Additional subjects were given formulations to try andquestionnaires to fill in, such that the sample size for each of thegroups rose to 102 subjects, of which:

-   -   82% reported less painful spots    -   80% said they had less shiny skin    -   76% felt their blackheads reduced    -   75% reported less swollen spots    -   75% found they had no spot recurrence.

Formulation 4 was well tolerated with no AEs reported and no erythema atthe test site.

This research demonstrates that Formulation 4 is a very effectivetreatment for acne. The reduction in sebum and both spots and blackheadsat 21 days also support the role of Formulation 4 in preventing futurerecurrence of the symptoms.

As mentioned, the objectively measured results demonstrated that againstthe control product the test products:

-   -   Significantly reduced surface sebum (by up to 50%);    -   Significantly reduced Comedones;    -   Significantly reduced the numbers of papules and pustules.

TABLE 4 summary of results Results of objective assessment Formulation4.2 Formulation 4 Reduction in sebum after. . . 1 week >20%; 1week >20%; 3 weeks >50% 3 weeks >50% Reduction in comedones after. . . 1week >40%; 1 week = 50%; 3 weeks >90% 3 weeks >80% Reduction in pustulesafter. . . 1 week = 90%; 1 week >90%; 3 weeks >98% 3 weeks >98% Resultsof subjective assessment: % agreeing/strongly agreeing Product reducedblackheads 64.7% 76.5% Product stopped spots recurring 69.6% 74.5% Spotsless painful (tender to touch) 66.7% 82.4% Spots not as swollen 73.5%75.5% Skin less shiny 59.8% 80.4% Skin less oily 67.6% 77.5% Skin lessgreasy 69.6% 75.5%

These results show the multifunctional nature of the formulation. It isbelieved that the colloidal particles in the formulations both assist inthe clearing of the excess sebum from the skin surface, whilst theirpenetration into the skin drags the chlorhexidine in solution into theskin structure, killing bacteria. The tethered zinc may also have had arole in both reducing sebum production and/or having an anti-microbialfunction.

Methods

TABLE 5 Summary Protocol Title: An in-use study in healthy volunteers toinvestigate the anti-spot efficacy of two test formulations, usingobjective instrumental assessments of skin sebum levels and subjectivevisual assessments of lesion prevalence, against a placebo following a3-week use period. Study design: Single-blind, within-subjectcomparison. Test Groups: Formulation 4.2. CBL-DERM-14-005-E Formulation4. CBL-DERM-006-F The two test formulations were identical apart fromcontaining colloidal particles that had slightly different SPC:Tweenratios Control: placebo. CBL-DERM-14-007/8-P Dose regime: The testgroups followed a 12-week usage schedule according to treatment specificin-use regimes. Duration of 21 Days. study: Number of 36 subjectscompleted the active phase for group 1, 37 subjects for subjects: group2 and 20 subjects completed assessments for group 3. Duration of StudyStarted: w/c 26, Jan. 2015 study: Study Ended: w/e 20, Feb. 2015Location: Princeton Consumer Research Ltd. 307 College Road EastPrinceton New Jersey 08540

Test Formulations

1. FORMULATION 4.2 AND FORMULATION 4 (CBL-DERM-14-005-E andCBL-DERM-006-F)

SOY PHOSPHATIDYLCHOLINE

BUTYLHYDROXYANISOLE

ETHANOL

ZINC STEARATE

METHYL-4-HYDROXYBENZOATE

ETHYL-4-HYDROXYBENZOATE

BENZYLALCOHOL

POLYSORBATE 80

CHLORHEXIDINE DIGLUCONATE

CITRIC ACID MONOHYDRATE

DI-SODIUM HYDROGEN ORTHOPHOSPHATE ANHYDROUS

SODIUM HYDROXIDE

CARBOPOL 974P

GLYCEROL

Linalool

WATER

The quantity of each component used is shown in the tables below.Formulation 4 (CBL-DERM-14-006-F) was the same as the exemplarycomposition set out in Example Formulation 4, above, except thatFormulation 4 (CBL-DERM-14-006-F) contains carbopol instead of methylcellulose (at the same quantity) and 0.11300 g of sodium hydroxide (theamount of water in the gel phase is adjusted accordingly).

2. CONTROL (CBL-DERM-14-007/8-P)

METHYL-4-HYDROXYBENZOATE

ETHYL-4-HYDROXYBENZOATE

CITRIC ACID MONOHYDRATE

DI-SODIUM HYDROGEN ORTHOPHOSPHATE DODECAHYDRATE

SODIUM HYDROXIDE

CARBOPOL 974P (CARBOMER)

WATER

Summary Formulation Information

Summary of Formulation 4 (CBL-DERM-14-006-F)

Quantity Required Per Ingredient 100 g Final Product (g) SPC (dry mass)6.870 Polysorbate 80 0.850 Benzylalcohol 0.525 Methyl-4-hydroxybenzoate0.250 Ethyl-4-hydroxybenzoate 0.250 Butylhydroxyanisol 0.020 Linalool0.100 Disodium hydrogen phosphate 12 H₂O 0.530 Citric acid monohydrate0.128 Glycerol 3.000 Ethanol 3.569 Sodium hydroxide 0.113 Carbopol 974PNF 0.750 Water 81.463 Agent of interest - tethered to Tethersomes Zincstearate 0.082 Agent of interest - present in continuous phaseChlorhexidine digluconate (20% solution) ** 1.5 pH 5.5 ** % with respectto the salt

Summary of Formulation 4.2 (CBL-DERM-14-005-E)

Quantity Required Per Ingredient 100 g Final Product (g) SPC (dry mass)7.146 Polysorbate 80 0.472 Benzylalcohol 0.525 Methyl-4-hydroxybenzoate0.250 Ethyl-4-hydroxybenzoate 0.250 Butylhydroxyanisol 0.020 Linalool0.100 Disodium hydrogen phosphate 12 H₂O 0.530 Citric acid monohydrate0.128 Glycerol 3.000 Ethanol 3.418 Sodium hydroxide 0.113 Carbopol 974PNF 0.750 Water 81.716 Agent of interest - tethered to Tethersomes Zincstearate 0.082 Agent of interest - present in continuous phaseChlorhexidine digluconate (20% solution) ** 1.5 pH 5.5 ** % with respectto the salt

Summary of Control (CBL-DERM-14-007/8-P)

Quantity Required Per Ingredient 100 g Final Product (g) SPC (dry mass)Polysorbate 80 Benzylalcohol Methyl-4-hydroxybenzoate 0.250Ethyl-4-hydroxybenzoate 0.250 Butylhydroxyanisol Linalool Disodiumhydrogen phosphate 12 H₂O 0.530 Citric acid monohydrate 0.128 GlycerolEthanol Sodium hydroxide 0.150 Carbopol 974P NF 1.000 Water 97.692 Agentof interest - tethered to Tethersomes Zinc stearate Agent of interest -present in continuous phase Chlorhexidine digluconate (20% solution) **pH 5.5 ** % with respect to the salt

Detailed Formulation Information

Formulation 4 (CBL-DERM-14-006-F)

Quantity Required Per 100 g Final Product (g) Organic Phase - EmptySequessome Intermediate SPC 3.43500 Ethanol 1.82550 BHA 0.01000Methyl-4-hydroxybenzoate 0.12500 Ethyl-4-hydroxybenzoate 0.12500 Benzylalcohol 0.26250 Polysorbate 80 0.42500 Linalool 0.05000 Organic PhaseSub-Total 6.25800 Aqueous Phase - Empty Sequessome Intermediate Citricacid monohydrate 0.03600 diNa hydrogen phos 12H2O 0.14400 Water 22.72850Aqueous Phase Sub-Total 22.90850 Empty Sequessome Intermediate Total29.16650 Organic Phase - Zinc Stearate Intermediate SPC 3.43500 Ethanol1.74350 BHA 0.01000 Methyl-4-hydroxybenzoate 0.12500Ethyl-4-hydroxybenzoate 0.12500 Benzyl alcohol 0.26250 Polysorbate 800.42500 Zinc stearate 0.08200 Linalool 0.05000 Organic Phase Sub-Total6.25800 Aqueous Phase - Zinc Stearate Intermediate Citric acidmonohydrate 0.03600 diNa hydrogen phos 12H2O 0.14400 Water 22.72850Aqueous Phase Sub-Total 22.90850 Zinc Tethersome Intermediate Total29.16650 Sum of Sequessome and Tethersome 58.33300 Intermediates GelPhase - Final Product Citric acid monohydrate 0.05600 diNa hydrogen phos12H2O 0.24200 Sodium hydroxide 0.11300 Carbopol 974P 0.75000 Glycerol3.00000 Water 36.00600 Gel Phase Sub-Total 40.16700 Chlorhexidinedigluconate 1.50000 Overall Total 100.00000

Formulation 4.2 (CBL-DERM-14-005-E)

Quantity Required Per 100 g Final Product (g) Organic Phase - EmptySequessome Intermediate SPC 3.57300 Ethanol 1.75000 BHA 0.01000Methyl-4-hydroxybenzoate 0.12500 Ethyl-4-hydroxybenzoate 0.12500 Benzylalcohol 0.26250 Polysorbate 80 0.23600 Linalool 0.05000 Organic PhaseSub-Total 6.13150 Aqueous Phase - Empty Sequessome Intermediate Citricacid monohydrate 0.06400 diNa hydrogen phos 12H2O 0.26500 Water 34.30000Aqueous Phase Sub-Total 34.62900 Empty Sequessome Intermediate Total40.76050 Organic Phase - Zinc Stearate Intermediate SPC 3.57300 Ethanol1.66800 BHA 0.01000 Methyl-4-hydroxybenzoate 0.12500Ethyl-4-hydroxybenzoate 0.12500 Benzyl alcohol 0.26250 Polysorbate 800.23600 Zinc stearate 0.08200 Linalool 0.05000 Organic Phase Sub-Total6.13150 Aqueous Phase - Zinc Stearate Intermediate Citric acidmonohydrate 0.06400 diNa hydrogen phos 12H2O 0.26500 Water 34.30000Aqueous Phase Sub-Total 34.62900 Zinc Tethersome Intermediate Total40.76050 Sum of Sequessome and Tethersome 81.52100 Intermediates GelPhase - Final Product Sodium hydroxide 0.11300 Carbopol 974P 0.75000Glycerol 3.00000 Water 13.11600 Gel Phase Sub-Total 16.97900Chlorhexidine digluconate 1.50000 Overall Total 100.00000

Results

Objectively Measured Parameters

Sebum Reduction

TABLE 6 Average reduction in Sebum score from Day 0. A negative scoreindicates an increase in sebum. n Day 3 Day 7 Day 14 Day 21 Formulation36 20.47 34.94 71.64 90.11 4.2 Formulation 4 37 14.51 36.30 85.35 98.49Control 20 −21.30 −48.60 −40.35 −44.95 1 vs. 2 t test p- 0.0163 0.67670.2065 0.3936 1 vs. Control values 7.04E−16 5.27E−23 9.74E−11 5.69E−16 2vs. Control 1.06E−14 1.62E−24 1.10E−14 4.63E−17

At all time points, both test formulations reduced sebum significantlymore than the control at p<0.1%. On day 3 Formulation 4.2 was betterthan Formulation 4 at p<0.02. There was no significant difference in theeffectiveness of the two test formulations at all later time points.

By Day 21, both test formulations had reduced the average sebum levelsto less than 50% of the starting value (see FIG. 4).

Comedone Reduction

TABLE 7 Average reduction in occurrence of comedones from Day 0. Apositive score indicates an increase in comedones. n Day 3 Day 7 Day 14Day 21 Formulation 36 −1.44 −2.31 −2.92 −3.44 4.2 Formulation 4 37 −1.14−1.97 −2.84 −3.38 Control 20 1.20 1.30 1.25 0.35 1 vs. 2 t test p- 0.4140.468 0.903 0.934 1 vs. Control values 2.35E−06 2.98E−06 4.66E−056.63E−04 2 vs. Control 1.08E−07 2.93E−05 7.20E−05 8.97E−05

At all time points, both test formulations reduced comedonessignificantly more than the control at p<0.1%. There was no significantdifference in the effectiveness of the two test formulations at all timepoints (see FIG. 5).

Papule Reduction

TABLE 8 Analysis of progression participants who had blemishes at day 0n Day 0 Day 3 Day 7 Day 14 Day 21 Formulation 4.2 5 13 5 1 0 0Formulation 4 7 16 9 5 0 0 Control 4 10 9 8 5 3

In those participants who began the trial with at least one papule, alltreatments showed a reduction in papule numbers over 21 days.

TABLE 9 Probabilities of U-statistic (Mann-Whitney) calculated on lesionreductions from Day 0 D3 v D7 vs D14 vs D21 vs D0 D0 D0 D0 1 vs. 2 0.1460.044 0.373 0.373 1 vs. Control 0.043 0.019 0.056 0.089 2 vs. Control0.110 0.093 0.110 0.254

The test statistic (U) shows that Formulation 4.2 was better thanControl @<5% on Day 3 and Day 7 and @<10% on Day 14 and Day 21 andbetter that Formulation 4 @<5% on Day 7. Formulation 4 was better thanControl @<10% on Day 7.

Pustule Reduction

TABLE 10 Reduction in pustule numbers from day 0. A positive scoreindicates an increase in numbers of pustules. n Day 3 Day 7 Day 14 Day21 Formulation 36 −1.28 −1.97 −2.11 −2.17 4.2 Formulation 4 37 −0.92−1.49 −1.57 −1.59 Control 20 0.60 0.05 0.15 −0.75 1 vs. 2 t test p-0.127 0.221 0.228 0.226 1 vs. Control values 2.71E−08 2.00E−05 9.67E−051.37E−02 2 vs. Control 8.60E−07 7.48E−04 1.04E−03 9.15E−02

At all time points from Day 3 to Day 14, both test formulations reducedpustule numbers significantly more than the control at p<0.1%. At Day21, Formulation 4.2 still outperformed control at p<2.0%; Formulation 4outperformed control at p<10%. There was no significant difference inthe effectiveness of the two test formulations at all time points (seeFIG. 6).

REFERENCES

1. Janiczek-Dolphin N1, Cook J, Thiboutot D, Harness J, Clucas A: Cansebum reduction predict acne outcome? Br J Dermatol. 2010Oct;163(4):683-8. doi: 10.1111/j. 1365-2133.2010.09878

Example 4: Anti-Ageing Trials. A Summary of Trials and Results SUMMARY

Two trials of novel products have been undertaken to assess theirefficacy and how they were perceived and received by users. See FIGS. 7,8, 9 and 10.

PROAWR1: A home-use study demonstrating the subjective and objectiveimprovements in wrinkles and skin appearance after treatment with twodifferent formulations of the same ingredients used with three differentregimes

PROAWR2: home-use study in three parallel groups of a preferredformulation and regime against a marketed comparator (in-market productA) and a control (placebo) product.

The test formulas in the two trials were variants of the sameingredients, formulated by blending a mixture of Sequessome-basedvesicles (i.e. colloidal particles), some of which had additionalmoieties tethered to the vesicle membrane.

Headline Results

PROAWR1:

-   -   100% of all participants on all three regimes had measured        reduction in wrinkle volume after the first week    -   The average scores both wrinkle volume reduction from baseline        measurements and the reduction in appearance of wrinkles from        baseline scores were highly significant (p<0.001) from week one    -   There was mean reduction in actual wrinkle volume of 17.46% at        week 12; the mean Glogau (appearance) score had reduced by        25.86% over the same period    -   In the subjective assessments:        -   100% reported that their skin felt smoother and healthier            and looked and felt plumper after 8 weeks        -   Over 90% reported that their skin looked smoother and            healthier, felt more elastic and that the bags and circles            under their eyes had reduced        -   80.8% of participants claimed that after 8 weeks their skin            looked younger by between 5 to 15 years; 50% claimed their            skin looked younger by between 10 to 15 years.

PROAWR2:

For all three independently assessed metrics (actual wrinkle volume, theappearance of fine lines and wrinkles and the appearance of peri-orbitaldark circles):

-   -   Both the Test Formulation and the Commercial Product (in-market        product A) were significantly better than the control in all        three measures—no improvements were seen for the control group.    -   The Test Formulation was equally effective as the in-market        Commercial Product (in-market product A).

PROAWR1: A home-use study demonstrating the subjective and objectiveimprovements in wrinkles and skin appearance after treatment with aformulation in accordance with the present invention in healthy subjectswith ageing and thinning skin

SUMMARY

This trial was conducted with two anti-ageing formulations in accordancewith Example Formulation 1, “Dilute” and “Concentrated”. In theConcentrated version, the tethered ascorbate and tethered peptides werepresent at twice the concentration that they were in the Dilute version.

The aim of this initial study was to examine the effectiveness ofincluding these tethered moieties and whether the concentration of themin the formulations and the regime with which they were applied wouldaffect their objective performance and the subjective perception of themby the participants.

Three regimes were followed:

-   -   Dilute: Applied twice a day all over the face and neck    -   Concentrated:Applied once a day to targeted lines and wrinkles    -   Combined: Participants on this regime used both products, as        directed above

The results showed that for the combined treatment:

-   -   100% of all participants on all three regimes had measured        reduction in wrinkle volume after the first week.    -   The average scores both wrinkle volume reduction from baseline        measurements and the reduction in appearance of wrinkles from        baseline scores were highly significant (p<0.001) from week one.    -   There was mean reduction in actual wrinkle volume of 17.46% at        week 12; the mean Glogau (appearance) score had reduced by        25.86% over the same period.    -   In the subjective assessments:        -   100% reported that their skin felt smoother and healthier            and looked and felt plumper after 8 weeks        -   Over 90% reported that their skin looked smoother and            healthier, felt more elastic and that the bags and circles            under their eyes had reduced        -   80.8% of participants claimed that after 8 weeks their skin            looked younger by between 5 to 15 years; 50% claimed their            skin looked younger by between 10 to 15 years.

TABLE 11 Summary Protocol Title: A randomised home-use study in threeparallel groups of 30 healthy volunteers, with wrinkles, to assess theefficacy of two anti-ageing products and one anti-ageing regime. Studydesign: Single-blind, within-subject comparison, whole face design. TestGroups: Group Test product Regime Group 1. DH3943 Used twice a day,morning and evening, in an n = 26 even layer to the face and neck Group2. DH3942 Apply once a day in the evening to targeted n = 27 lines orwrinkles Group 3. DH3943 Apply DH 3943 twice a day in an even layer to n= 26 and the face and neck DH3942 Once a day apply a small quantity ofDH3942 to targeted lines or wrinkles. Duration of 12 week treatmentperiod, with assessments at Baseline, Weeks 1, 2, 4, study: 6, 8 and 12.Study Started: w/c 11, Aug. 2014 Study Ended: w/e 21, Nov. 2014 Numberof 79 subjects completed the study up to Week 8. 62 subjects completedsubjects: the study including the 4 week extension to week 12. Method:Subjects underwent Profilometry assessments after which they were issuedwith an instruction sheet, diary cards and the test article on Week 0(baseline). Subjects returned to the Test Centre on Weeks 1, 2, 4, 6 and8 for Profilometry assessments. Visual assessments were performed onboth ageing and peri-orbital dark circles at baseline and Weeks 1, 2, 4,6 and 8. The subjects completed SPQ's at Week 4 and Week 8. DIC (DigitalImage Capture) was performed on approximately 40% of subjects in eachgroup at baseline, Week 2, 4 and 8). A 4 week extension was authorisedfor the study in which groups 1 and 2 were instructed to continue use ofthe product and undergo further profilometery and visual assessments atweek 12. 50% of Group 3 were instructed to continue use of the regimenwhilst the remaining 50% were instructed to use a bland moisturiser.Following week 12 assessments subjects were compensated for their timeand inconvenience and exited from the study. Location: PrincetonConsumer Research Ltd. Harbour House 23 Chandlers Quay Maldon CM9 4LFUnited Kingdom

TABLE 12 Test Products DH3942 - DH3943 - “Dilute” “Concentrated” SoyPhosphatidylcholine Soy Phosphatidylcholine ButyIhydroxytoIueneButylhydroxytoluene Sodium metabisulphite Sodium metabisulphite Citricacid Citric acid di-Sodium hydrogen di-Sodium hydrogen phosphatedodecahydrate phosphate dodecahydrate Sodium EDTA Sodium EDTAMethyl-4-hydroxybenzoate Methyl-4-hydroxybenzoateEthyl-4-hydroxybenzoate Ethyl-4-hydroxybenzoate BenzylalcoholBenzylalcohol Polysorbate 80 Polysorbate 80 Sodium hydroxide Sodiumhydroxide Carbopol 974P Carbopol 974P Glycerol Glycerol Ethanol EthanolWater Water Ascorbyl Palmitate Ascorbyl Palmitate* PalmitoylTripeptide-1 Palmitoyl Tripeptide-1* Palmitoyl Tetrapeptide-7 PalmitoylTetrapeptide-7* Linalool Linalool *In DH3942, these three componentswere present at double the concentration that they were in DH3943.DH3942 is the same as that in the exemplary composition set out inExample Formulation 1, above.

Example Formulation 1, above.

Results

Effect on Wrinkle Volume

100% of participants experienced reductions in wrinkle volume.

TABLE 13 Average wrinkle volume at time t, expressed as a percentage ofaverage volume at time 0 (=100%). Standard deviation in brackets. Pvalue is the result of a t test comparison to the Week 0 data. Weeks 1 24 6 8 12 DH3943 - 96.9% 93.8% 91.9% 90.5% 89.0% “Dilute” (1.3%) (1.6%)(1.9%) (1.9%) (2.1%) p < 0.001 p < 0.001 p < 0.001 p < 0.001 p < 0.001DH3942 - 96.3% 93.7% 90.8% 88.8% 87.1% “Concentrated” (2.1%) (2.7%)(2.7%) (2.7%) (2.7%) p < 0.001 p < 0.001 p < 0.001 p < 0.001 p < 0.001Dilute + 96.5% 92.7% 88.9% 85.2% 82.3% 83% Concentrated (0.9%) (1.1%)(1.5%) (1.7%) (1.9%) (1.9%) p < 0.001 p < 0.001 p < 0.001 p < 0.001 p <0.001 p < 0.001

Effect on Glogau Score (Appearance of Fine Lines and Wrinkles)

Glogau scores are assessments made by an independent assessor of theseverity of appearance of lines and wrinkles.

TABLE 14 Average Glogau score at time t, expressed as a percentage ofaverage volume at time 0 (=100%). Standard deviation in brackets. Pvalue is the result of a t test comparison to the Week 0 data. Weeks 1 24 6 8 12 DH3943 - 99.2%  100% 99.6% 95.4% 88.8% “Dilute” (3.9%) (0.0%)(2.0%) (7.0%)  (8.9%) p = 0.163 N/A p = 0.163 p = 0.001 p < 0.001DH3942 - 99.2% 98.8% 96.5% 94.7% 93.1% “Concentrated” (4.3%) (4.3%)(7.3%) (8.9%) (12.0%) p = 0.163 p = 0.081 p = 0.008 p = 0.001 p = 0.002Dilute + 95.7% 91.8% 87.7% 86.5% 81.5% 73.8% Concentrated (6.3%) (7.7%)(7.6%) (8.6%) (10.0%) (10.4%) p < 0.001 p < 0.001 p < 0.001 p < 0.001 p< 0.001 p < 0.001

Subjective Questionnaire Results

TABLE 15 % of subjects Agreeing/Strongly agreeing with the statement DH3942 - Conc DH 3943 - Dil Combined Week Week Week Week Week WeekStatement 4 8 4 8 4 8 Felt smoother 23.1% 50.0% 30.3% 81.5% 46.2% 100.0%Looked smoother 26.9% 61.5% 55.6% 70.4% 30.8% 96.2% Skin tone more even50.0% 92.3% 29.6% 74.1% 15.4% 88.5% Acne scarring reduced 19.2% 84.6%37.0% 55.6% 11.5% 38.5% Blemishes reduced 26.9% 73.1% 29.6% 100.0% 3.9%80.8% Complexion improved 30.8% 61.5% 22.2% 100.0% 7.7% 76.9% Complexionmore radiant 19.2% 38.5% 29.6% 96.3% 23.1% 88.5% Age spots/hyperpigmentation reduced 26.9% 53.9% 55.6% 55.6% 61.5% 88.5% Felt more firm38.5% 50.0% 55.6% 92.6% 42.3% 80.8% Looked more firm 19.2% 53.9% 22.2%40.7% 34.6% 100.0% Felt more plump 30.8% 57.7% 40.7% 100.0% 19.2% 100.0%Looked more plump 19.2% 46.2% 59.3% 92.6% 34.6% 100.0% Complexion lookedhealthier 42.3% 76.9% 25.9% 92.6% 23.1% 96.2% Skin feels healthier 53.9%61.5% 74.1% 88.9% 42.3% 100.0 Skin felt more elastic 46.2% 73.1% 55.6%70.4% 26.9% 92.3% Bags/circles reduced 30.8% 57.7% 44.4% 96.3% 30.8%92.3% Most effective anti-ageing used 42.3% 76.9% 55.6% 81.5% 30.8%76.9% Would use instead of current 46.2% 69.2% 55.6% 92.6% 65.4% 92.3%Easy to use alongside current products 46.2% 80.8% 100.0% 96.3% 34.6%61.5% Would recommend to a friend 65.4% 76.9% 81.5% 100.0% 73.1% 100.0%Would buy the product 38.5% 76.9% 18.5% 81.5% 84.6% 92.3% Skin lookedyounger by 20+ years 0.0% 0.0% 0.0% 0.0% 0.0% 0.0% 15+ years 0.0% 3.9%0.0% 3.7% 0.0% 7.7% 10+ years 3.9% 19.2% 7.4% 22.2% 3.9% 42.3% 5+ years23.1% 42.3% 81.5% 55.6% 42.3% 30.8% Same 73.1% 34.6% 11.1% 18.5% 50.0%19.2%

PROAWR2: A randomised home-use study in three parallel groups of 30healthy volunteers, with wrinkles, to assess the efficacy of oneanti-ageing product against a placebo and a comparator with anadditional 73 subjects to complete a subjective user-trial using onlythe product

SUMMARY

This trial tested a preferred test anti-ageing formulation in accordancewith the present invention (the “Concentrated” formulas from PROAWR1),against an in-market anti-ageing Commercial Product (in-market productA) and a control formulation.

The aim of this study was to test:

-   -   a) That previous results seen with the preferred formulation        were real (control comparison)    -   b) To assess the effectiveness against a currently marketed        formulation for anti-ageing.

The results showed that:

For all three independently assessed metrics (actual wrinkle volume, theappearance of fine lines and wrinkles and the appearance of peri-orbitaldark circles):

-   -   Both the Test Formulation and the Commercial Product were        significantly better than the control in all three measures—no        improvements were seen for the control group.    -   The Test Formulation was equally effective as the in-market        Commercial Product.

For the subjective assessment of the products:

-   -   Both the Test Formulation and the Commercial Product were        significantly better than the control in all measures    -   The Test Formulation was rated higher than the Commercial        Product for:        -   Skin looked smoother        -   Skin felt more plump        -   Skin felt healthier        -   Skin felt more elastic        -   Complexion looked healthier        -   Skin tone looked more even        -   Eye puffiness reduction        -   Effectiveness in anti-ageing        -   Lifted sagging skin        -   Hydration of skin        -   Reducing pore size

TABLE 16 Summary Protocol Title: A randomised home-use study in threeparallel groups of 30 healthy volunteers, with wrinkles, to assess theefficacy of one anti-ageing product against a placebo and a comparatorwith an additional 70 subjects to complete a subjective user-trial usingonly the product. Study design: Single-blind, within-subject comparison,whole face design. Test Groups: Group Test product Regime Group 1. TestFormulation Apply twice a day Group 2. Control Apply all over to Group3. In-market product A face and neck Apply additional amounts to areaswhere wrinkles/ageing is more noticeable Apply a thin layer to thesub-orbital eye sacks Duration of 8 week treatment period, withassessments at Baseline, Week 4, and study: Week 8. Study Started: w/c16, Feb. 2015 Study Ended: w/e 24, Apr. 2015 Method: Subjects underwentProfilometry assessments after which they were issued with aninstruction sheet, diary cards and the test article on Week 0(baseline). Subjects returned to the Test Centre on Weeks 1, 2, 4, 6 and8 for Profilometry assessments. When the subjects returned to the TestCentre on Week 8, they returned any unused test articles and theircompleted diary cards. Visual assessments were performed on both ageingand peri-orbital dark circles at baseline and Weeks 1, 2, 4, 6 and 8.The subjects completed SPQ's at Week 8. Professional photography wasperformed on approximately 40% of subjects in each group at baseline,and Weeks 1, 2, 4 and 8 with as many subjects as possible for group 1being photographed above and beyond 40%. 70 subjects were also issuedthe product to use at home for the duration of the test period andcompleted an SPQ at the same time points as the active groups. Location:Princeton Consumer Research Ltd. Harbour House 23 Chandlers Quay MaldonCM9 4LF United Kingdom

TABLE 17 Test Products Test Formulation Control Soy PhosphatidylcholineButyIhydroxytoIuene Ethanol Methyl-4-hydroxybenzoateMethyl-4-hydroxybenzoate Ethyl-4-hydroxybenzoate Ethyl-4-hydroxybenzoateBenzylalcohol Polysorbate 80 Linalool Ascorbyl Palmitate PalmitoylTripeptide-1 Palmitoyl Tetrapeptide-7 Sodium metabisulphite Citric acidmonohydrate Citric acid monohydrate di-Sodium hydrogen di-Sodiumhydrogen orthophosphate anhydrous orthophosphate anhydrous Sodium EDTASodium hydroxide Sodium hydroxide Carbopol 974P Carbopol 974P GlycerolWater Water

Results

Effect on Wrinkle Volume

TABLE 18 Average wrinkle volume at time t, expressed as a percentage ofaverage volume at time 0 (= 100%). Standard deviation in brackets. Pvalue is the result of a t test comparison to the Week 0 data. n Week 1Week 2 Week 4 Week 6 Week 8 Test 27 95.9% 93.1% 90.7% 88.8% 87.5%Formulation (2.0%) (2.9%) (3.6%) (3.9%) (4.1%) p < 0.001 p < 0.001 p <0.001 p < 0.001 p < 0.001 Control 28 100.2%  100.5%  100.3%  100.4% 100.5%  (1.0%) (2.0%) (2.9%) (3.0%) (3.3%) P = 0.31  P = 0.17  P = 0.57 P = 0.51  P = 0.39  In-market 29 95.8% 92.9% 90.5% 88.1% 86.2% product A(2.5%) (3.3%) (3.8%) (4.1%) (4.3%) p < 0.001 p < 0.001 p < 0.001 p <0.001 p < 0.001

TABLE 19 p values for t-tests between different treatments Week 1 Week 2Week 4 Week 6 Week 8 Test Formulation vs. p < 0.001 p < 0.001 p < 0.001p < 0.001 p < 0.001 Control Test Formulation vs. in- p = 0.554 p = 0.623p = 0.604 p = 0.817 p = 0.947 market product A In-market product A vs. p< 0.001 p < 0.001 p < 0.001 p < 0.001 p < 0.001 Control

-   -   Test Formulation and in-market product A significantly different        from Control.    -   No significant difference between Test Formulation and in-market        product A.

Effect on Glogau Score (Appearance of Fine Lines and Wrinkles)

Glogau scores are assessments made by an independent assessor of theseverity of appearance of lines and wrinkles.

TABLE 20 Average Glogau score at time t, expressed as a percentage ofaverage volume at time 0 (=100%). Standard deviation in brackets. Pvalue is the result of a t test comparison to the Week 0 data. n Week 1Week 2 Week 4 Week 6 Week 8 Test 27 92.6% 90.7% 79.5% 78.5% 73.9%Formulation (8.5%) (8.3%) (9.8%) (8.9%)  (7.4%) p < 0.001 p < 0.001 p <0.001 p < 0.001 p < 0.001 Control 28 101.7%  100.0%  102.8%  105.0% 103.9%  (10.8%)  (10.7%)  (8.0%) (13.4%)  (12.8%) p = 0.81  p = 0.79  p= 0.21  p = 0.12  p = 0.31  In-market 29 91.6% 88.9% 82.2% 79.9% 74.3%product A (8.1%) (8.4%) (8.5%) (9.5%) (10.5%) p < 0.001 p < 0.001 p <0.001 p < 0.001 p < 0.001

TABLE 21 p values for t-tests between different treatments Week 1 Week 2Week 4 Week 6 Week 8 Test Formulation vs. p = 0.266 p = 0.198 p = 0.004p < 0.001 p < 0.001 Control Test Formulation vs. in- p = 0.976 p = 0.998p = 0.681 p = 0.698 p = 0.799 market product A in-market product A vs. p= 0.318 p = 0.245 p = 0.018 p = 0.005 p = 0.001 Control

-   -   Test Formulation and in-market product A significantly different        from Control from week 4.    -   No significant difference between Test Formulation and in-market        product A.

Effect on Peri-Orbital Dark Circles

TABLE 22 Average PO Dark circle score at time t, expressed as apercentage of average volume at time 0 (=100%). Standard deviation inbrackets. P value is the result of a t test comparison to the Week 0data. n Week 1 Week 2 Week 4 Week 6 Week 8 Test 27 72.2% 48.1% 38.9%29.6% 27.8% Formulation (34.9%) (37.9%) (42.4%) (42.2%) (37.6%) p <0.001 p < 0.001 p < 0.001 p < 0.001 p < 0.001 Control 28  100% 112.5% 114.3%  119.6%  121.4%    (0%) (44.4%) (54.2%) (59.8%) (58.4%) p = N/A p= 0.49  p = 0.79  p = 0.63  p = 0.45  In-market 29 77.6% 56.9% 34.5%34.5% 31.0% product A (34.3%) (43.8%) (40.3%)  42.5%) (41.0%) p < 0.001p < 0.001 p < 0.001 p < 0.001 p < 0.001

TABLE 23 p values for t-tests between different treatments Week 1 Week 2Week 4 Week 6 Week 8 Test Formulation vs. p = 0.013 p < 0.001 p < 0.001p < 0.001 p < 0.001 Control Test Formulation vs. in- p = 0.949 p = 0.589p = 0.794 p = 0.562 p = 0.962 market product A in-market product A vs. p= 0.008 p < 0.001 p < 0.001 p < 0.001 p < 0.001 Control

-   -   Test Formulation and in-market product A significantly different        from Control from week 1.    -   No significant difference between Test Formulation and in-market        product A.

Subjective Questionnaire Results

TABLE 24 In market Test Product Formulation A Control N = 27 + 73* N =26 N = 28 After using the product. . . my skin felt significantlysmoother 82.00% 89.66% 28.57% my skin looked significantly smoother85.00% 55.17% 46.43% my skin felt significantly more firm 89.00% 89.66%42.86% my skin looked significantly more firm 54.00% 58.62% 46.43% myskin felt significantly more plump 86.21% 25.00% my skin lookedsignificantly more plump 56.00% 68.97% 25.00% my skin feelssignificantly healthier 86.00% 79.31% 46.43% my skin felt more elastic83.00% 82.76% 28.57% my complexion was visibly improved 58.00% 65.52%25.00% my complexion was visibly more radiant 61.00% 82.76% 21.43% mycomplexion looked significantly healthier 90.00% 86.21% 32.14% my skintone looked significantly more even 90.00% 79.31% 25.00% any agespots/hyperpigmentation were 67.00% 72.41% 46.43% significantly reducedthe bags under my eyes were significantly 89.00% 35.71% reduced thepuffiness all around my eyes was 88.00% 86.21% 39.29% significantlyreduced the dark circles under my eyes were 86.00% 89.66% 42.86%significantly reduced sagging skin was lifted 89.00% 86.21% 46.43% myskin looked: at least 5 years younger 76.00% 82.76% 39.29% at least 10years younger 35.00% 34.48%  0.00% The product. . . redefined saggedskin 58.00% 75.86% 28.57% has a pleasant fragrance 62.00% 58.62% 42.86%has a pleasant consistency and texture 79.31% 32.14% has a pleasantappearance 61.00% 65.52% 28.57% felt good on my skin after or upon66.00% 65.52% 32.14% application dried quickly 89.00% 32.14% was easy touse alongside my usual 86.21% 28.57% skincare products was the mosteffective anti-ageing serum I 86.00% 62.07% 39.29% have used redefinedfacial contours that were sagged 67.00% 72.41% 39.29% hydrated my skin86.21% 25.00% has visibly reduced the size of my pores 85.00% 68.97%17.86% has visibly reduced the appearance of any 69.00% 75.86% 25.00%thread veins I would. . . recommend the product to a friend 79.00%32.14% buy this product in place of my usual anti- 57.14% ageing serum*An additional 73 subjects used the Test Formulation and answered theself-assessment questionnaire but were not independently assessed forwrinkle volume, wrinkle appearance or appearance of peri-orbital darkcircles.

Example 5: A Study in Volunteers to Investigate the Sensation ProducedUsing a Formulation Comprising Capsaicin BACKGROUND

There are several products on the market that provide a warmingsensation when applied to the joint or muscle, to relieve aches orpains. These products contain chemicals such as capsaicin whichstimulate the nerve endings in the skin to create the warming sensations(without actually being related to a physical temperature change).

The inventors have developed a capsaicin-containing formulation inaccordance with the present invention, so that a warming sensation isassociated with the product, either at application or sometime after.The formulation provides a combined effect of the joint-pain relievingproperty of drug-free colloidal dispersions, combined with a warmingfeeling.

The formulation (TP2) comprised insoluble and inflexible liposomescomprising capsaicin and menthol in combination with Transfersomes™comprising menthol.

Transfersomes™/Sequessomes™/Tethersomes are flexible, deformablecolloidal particles that make their own way into the skin, whereasliposomes are rigid vesicles that would not ordinarily be expected topenetrate the skin by themselves unless some other force is applied(e.g. Sequessomes™/Transfersomes™/Tethersomes in the same mixtureopening pores and pulling/pushing the liposomes through).

The starting material used to make TP2 is set out below.

TP2 (PD-14-0073) was formed by taking 90 g of Start Material 1 andadding/mixing in 10 g of Start Material 2.

Start Material 1 (Flexible, Empty, Menthol-Fragranced Vesicles):

Quantity Required Per 100 g Final Product (g) Organic Phase - EmptyTransfersome Intermediate SPC (Dry Mass) 6.87000 Ethanol 3.65100 BHT0.02000 Methyl-4-hydroxybenzoate 0.25000 Ethyl-4-hydroxybenzoate 0.25000Benzyl alcohol 0.52500 Polysorbate 80 0.85000 Capsaicin 0.00000 Menthol0.10000 Organic Phase Sub-Total 12.51600 Aqueous Phase - EmptyTransfersome Intermediate Sodium dihydrogen orthophosphate 2 H₂O 0.03700disodium hydrogen orthophosphate 12 H₂O 0.45300 Sodium EDTA 0.10000Water 45.22700 Aqueous Phase Sub-Total 45.81700 IntermediateTransfersome Total 58.33300 Gel Phase - Final Product Sodium dihydrogenorthophosphate 2 H₂O 0.02400 disodium hydrogen orthophosphate 12 H₂O0.30200 Sodium hydroxide 0.63000 Carbopol 974P NF 1.25000 Glycerol3.00000 Water 36.46100 Gel Phase Sub-Total 41.66700 Overall Total100.00000

Start Material 2 (Capsaisin Liposomes):

Quantity Required Per 100 a Final Product (g) Organic Phase - LiposomeIntermediate SPC (Dry Mass) 6.87000 Ethanol 4.25100 BHT 0.02000Methyl-4-hydroxybenzoate 0.25000 Ethyl-4-hydroxybenzoate 0.25000 Benzylalcohol 0.52500 Polysorbate 80 0.00000 Capsaicin 0.25000 Menthol 0.10000Organic Phase Sub-Total 12.51600 Aqueous Phase - Liposome IntermediateSodium dihydrogen orthophosphate 2 H₂O 0.03700 disodium hydrogenorthophosphate 12 H₂O 0.45300 Sodium EDTA 0.10000 Water 45.22700 AqueousPhase Sub-Total 45.81700 Liposome Intermediate Total 58.33300 GelPhase - Final Product Sodium dihydrogen orthophosphate 2 H₂O 0.02400disodium hydrogen orthophosphate 12 H₂O 0.30200 Sodium hydroxide 0.63000Carbopol 974P NF 1.25000 Glycerol 3.00000 Water 36.46100 Gel PhaseSub-Total 41.66700 Overall Total 100.00000

Summary Formulae:

Title PD-14-0073 (90% Start Material 1 plus 10% Start Ingredient StartMaterial 1 Start Material 2 Material 2 SPC (dry mass) 6.870 g 6.870 g6.870 g Polysorbate 80 0.850 g — 0.765 g Benzylalcohol 0.525 g 0.525 g0.525 g Methyl-4-hydroxybenzoate 0.250 g 0.250 g 0.250 gEthyl-4-hydroxybenzoate 0.250 g 0.250 g 0.250 g Butylhydroxytoluene0.020 g 0.020 g 0.020 g Disodium EDTA 0.100 g 0.100 g 0.100 g Disodiumhydrogen phosphate 0.755 g 0.755 g 0.755 g 12 H₂O Sodium dihydrogenphosphate 2 0.061 g 0.061 g 0.061 g H₂O Glycerol 3.000 g 3.000 g 3.000 gEthanol 3.651 g 4.251 g 3.711 g Sodium hydroxide 0.630 g 0.630 g 0.630 gCarbopol 974P NF 1.250 g 1.250 g 1.250 g Water 81.688 g 81.688 g 81.688g Agent of interest - in continuous phase Capsaicin — 0.250 g 0.025 gAgent of interest - in Transfersome Menthol 0.100 g 0.100 g 0.100 gTotal mass 100.000 g 100.000 g 100.000 g

Results

The formulation was tried by three individuals. The results arepresented in the table below.

TABLE 26 Test product TP2 (PD-14-0073) Capsaicin In liposomes Menthol Inmembranes of Transfersomes ™and of liposomes CAPSAICIN: Onset ofsensation 5 to 30 mins Onset of optimal sensation 15 to 60 mins Durationof sensation 45 to 180 mins Intensity of sensation Pleasant MENTHOL:Duration of sensation 15 to 30 mins Intensity of sensation Enough

Conclusion/Summary

These early results are useful in showing that:

-   -   The warming sensation of capsaicin can be felt:        -   In Transfersomes™—in the same “phase” as a joint-targeting            colloidal particle        -   In liposomes—separate “phase” to the joint-targeting            colloidal particles

Example 6: A User Trial Study in Healthy Volunteers to Investigate theEfficacy of a Formulation Comprising Caffeine in Improving theAppearance of Periorbital Skin

This study tested the efficacy of a formulation comprising caffeine andtocopherol in the continuous phase and three types of deformablecolloidal particles: 1) Tethersomes to which palmitoyl ascorbate istethered, 2) Tethersomes to which palmitoyl tripeptide-1 is tethered,and 3) Tethersomes to which palmitoyl tetrapeptide-7 is tethered.

Methods

Summary Protocol

Study desiqn: Sinqle-blind Test article: CBL-DERM-15-013 Periorbitalskin serum Duration of treatment: 4 weeks Number of subjects: 102 Typeof subjects: Healthy male and female subjects (from a breadth ofethnicities) aged between 40 and 70 years old (in equal proportions, 33%each 10 year age group), from a breadth of ethnicities and with avariety of skin types (normal, combination, dry, oily, sensitive) whosuffer from the appearance of two of the following characteristics:Periorbital puffiness - swelling around the eyes Puffiness below thelower eyelids - ‘eye bags’ Periorbital dark circles caused by a) agingb) heredity and c) hyperpigmentation (dark skinned people) (There mustbe an equal representation of all three characteristics to ensure claimsobjectivity). All subjects must have visible wrinkles, fine lines andcrow's feet around the eye area Observations: Subjects were asked toapply the product as per usage instructions. They were then asked tocomplete a Self- Perception Questionnaire (SPQ) after two weeks and atthe end of the study after four weeks. Treatment: Subjects were issuedwith samples of the test article and directions for use followingstandard in-use application regime. Location Princeton Consumer ResearchHarbour House 23 Chandlers Quay Maldon CM9 4LF United Kingdom

Formulation (CBL-DERM-15-013)

Quantity Required Per 100 g Final Product (g) Organic Phase - 1600 ppmPAA Intermediate Soy phosphatidylcholine (SPC) 4.12200 Ethanol 1.99660Butyl hydroxyanisole (BHA) 0.01200 Methyl-4-hydroxybenzoate 0.15000Ethyl-4-hydroxybenzoate 0.15000 Benzyl alcohol 0.31500 Polysorbate 800.40800 Palmitoyl ascorbic acid 0.09600 +/− alpha tocopherol 0.20000Linalool 0.06000 Organic Phase Sub-Total 7.50960 Aqueous Phase - 1600ppm PAA intermediate Sodium metabisulphite 0.03000 Citric acidmonohydrate 0.04320 Disodium EDTA 0.06000 Disodium hydrogenorthophosphate 0.17280 dodecahydrate Caffeine 0.05000 Water 27.13420Aqueous Phase Sub-Total 27.49020 PAA Tethersome Intermediate Total34.99980 Organic Phase - 300 ppm Tetrapeptide Intermediate Soyphosphatidylcholine (SPC) 1.37400 Ethanol 0.74120 Butylhydroxyanisole(BHA) 0.00400 Methyl-4-hydroxybenzoate 0.05000 Ethyl-4-hydroxybenzoate0.05000 Benzyl alcohol 0.10500 Polysorbate 80 0.15300 Palmitoyl peptide0.00600 Linalool 0.02000 Organic Phase Sub-Total 2.50320 Aqueous Phase -300 ppm Tetrapeptide Intermediate Sodium metabisulphite 0.01000 Citricacid monohydrate 0.01440 Disodium EDTA 0.02000 Disodium hydrogenorthophosphate 0.05760 dodecahydrate Water 9.06140 Aqueous PhaseSub-Total 9.16340 Tetrapeptide Tethersome 11.66660 Intermediate TotalOrganic Phase - 300 ppm Tripeptide Intermediate Soy phosphatidylcholine(SPC) 1.37400 Ethanol 0.74120 Butylhydroxyanisole (BHA) 0.00400Methyl-4-hydroxybenzoate 0.05000 Ethyl-4-hydroxybenzoate 0.05000 Benzylalcohol 0.10500 Polysorbate 80 0.15300 Palmitoyl peptide 0.00600Linalool 0.02000 Organic Phase Sub-Total 2.50320 Aqueous Phase - 300 ppmTripeptide Intermediate Sodium metabisulphite 0.01000 Citric acidmonohydrate 0.01440 Disodium EDTA 0.02000 Disodium hydrogenorthophosphate 0.05760 dodecahydrate Water 9.06140 Aqueous PhaseSub-Total 9.16340 Tripeptide Tethersome 11.66660 Intermediate Total Sumof Sequessome and 58.33300 Tethersome Intermediates Gel Phase - FinalProduct Sodium hydroxide 0.16000 Carbopol 974P NF 0.75000 Glycerol3.00000 Citric acid monohydrate 0.05600 Disodium hydrogen orthophosphate0.24200 dodecahydrate Water 37.45900 Gel Phase Sub-Total 41.66700Overall Total 100.00000

Summary Formulae of CBL-DERM-15-013

Periorbital Ingredient (percentage; g per 100 g) Skin SPC (dry mass)6.870 Polysorbate 80 0.714 Benzylalcohol 0.525 Methyl-4-hydroxybenzoate0.250 Ethyl-4-hydroxybenzoate 0.250 Butylhydroxyanisole 0.020 Linalool0.100 Sodium metabisulphite 0.050 Disodium EDTA 0.100 Disodium hydrogenphosphate 12 H₂O 0.530 Citric acid monohydrate 0.128 Glycerol 3.000Ethanol *** 3.479 Sodium hydroxide 0.160 Carbopol 974P NF 0.750 Water82.716  Agent of interest - tethered to Tethersomes Palmitoyltripeptide-1 0.006 Palmitoyl tetrapeptide-7 0.006 Palmitoyl ascorbate0.096 Agent of interest - in continuous phase Tocopherol 0.200 Caffeine0.050 pH 5.5*  *Final pH approximate; to be confirmed

Results

Within-treatment analysis (p<0.05) of periorbital clinical assessmentshows there to be a statistically significant reduction in wrinkles(15.69%), fine lines (31.22%), crow's feet (21.47%), puffiness (38.07%),dark circles (26.01%) and eye bags (38.94%) after 4 weeks of productusage.

The product performed statistically favourably over the 4 week study inthe majority of attributes under Clearcast guidelines of advertising.The product showed a preference or favourability in the majority ofattributes:

-   -   After using the product, 96.08% of users agreed, or strongly        agreed the product reduced the appearance of puffiness and        swelling around their eyes.    -   After using the product, 96.08% of users agreed, or strongly        agreed their skin felt and looked less thin.    -   After using the product, 93.14% of users agreed, or strongly        agreed their skin felt more elastic.    -   After using the product, 96.08% of users agreed, or strongly        agreed the product reduced the appearance of swelling and        puffiness below the lower eyelids (eye bags).    -   After using the product, 90.20% of users agreed, or strongly        agreed the product lifted sagged skin (reduction in skin        droopiness and sagging).    -   After using the product, 92.16% of users agreed, or strongly        agreed the product reduced the appearance of under eye dark        circles.    -   After using the product, 95.10% of users agreed, or strongly        agreed their skin looked and felt more firm.    -   After using the product, 90.20% of users agreed, or strongly        agreed there was a reduction in fine lines (inc. crow's feet)        around the eye area.    -   After using the product, 87.25% of users agreed, or strongly        agreed there was a reduction in deep wrinkles (inc. crow's feet)        around the eye area.    -   After using the product, 93.14% of users agreed, or strongly        agreed their skin tone looked more even.    -   After using the product, 90.20% of users agreed, or strongly        agreed their eye contor looked and felt tighter.    -   After using the product, 90.20% of users agreed, or strongly        agreed their eye contour looked more toned/lifted.    -   After using the product, 96.08% of users agreed, or strongly        agreed their eyes looked rested/less tired.    -   After using the product, 97.06% of users agreed, or strongly        agreed their eye skin was more hydrated.    -   After using the product, 95.10% of users agreed, or strongly        agreed their skin looked smoother.    -   After using the product, 99.02% of users agreed, or strongly        agreed the product was gentle and well tolerated.    -   After using the product, 75.49% of users stated, yes, they would        buy this product instead of their usual product.    -   After using the product, 86.27% of users stated, yes, they would        recommend this product to a friend.    -   After using the product, 46.08% of users stated, they looked at        least 5 years younger.

Example 7: A User Trial Study in Healthy Volunteers to Investigate theEfficacy of a Formulation Comprising Salicylate and Tocopherol inImproving Skin Tone

This study tested the efficacy of a formulation comprising tocopherol inthe continuous phase and two types of colloidal particles: 1)Tethersomes to which tridecyl salicylate is tethered and 2) Tethersomesto which palmitoyl ascorbate and tocopheryl linoleate are tethered.

Methods

Summary Protocol

Study design: Single-blind Test article: CBL-DERM-15-014 skin tone serumDuration of treatment: 6 weeks Number of subjects: 110 Type of subjects:A user trial study in 110 healthy male (20%) and female (80%) subjectsaged between 20 and 70 years old (equal proportions of each 10 year agegroup; 20's, 30's, 40's, 50's, 60's, 70's); from a breadth ofethnicities with a variety of skin types who suffer from one pronouncedor two out of the following six conditions; melasma or chloasma spots,uneven skin tone, age, sun or “liver” spots, freckles, post-inflammatoryhyperpigmentation or periorbital hyperpigmentation (dark circles).Observations: Subjects were asked to apply the product as per usageinstructions. They were asked to complete a Self-PerceptionQuestionnaire (SPQ) at the end of Weeks 3 and 6. Treatment: Subjectswere issued with samples of the test article and directions for usefollowing standard in-use application regime. Location PrincetonConsumer Research Harbour House 23 Chandlers Quay Maldon CM9 4LF UnitedKingdom

Formulation (CBL-DERM-15-014)

Quantity Required Per 100 g Final Product (g) Organic Phase - TridecylSalicylate Intermediate Soy phosphatidylcholine (SPC) 3.43500 Ethanol1.67550 Butylhydroxyanisole (BHA) 0.01000 Methyl-4-hydroxybenzoate0.12500 Ethyl-4-hydroxybenzoate 0.12500 Benzyl alcohol 0.26250Polysorbate 80 0.42500 Tridecyl Salicylate 0.10000 +/− alpha tocopherol0.05000 Linalool 0.05000 Organic Phase Sub-Total 6.25800 Aqueous Phase -Tridecyl Salicylate Intermediate Citric acid monohydrate 0.03600Disodium EDTA 0.05000 Sodium metabisulphite 0.02500 Disodium hydrogenorthophosphate 0.14400 dodecahydrate Water 22.65350 Aqueous PhaseSub-Total 22.90850 Tridecyl Salicylate Tethersome 29.16650 IntermediateTotal Organic Phase - PAA/Tocopheryl Linoleate Intermediate Soyphosphatidylcholine (SPC) 3.43500 Ethanol 1.57950 Butylhydroxyanisole(BHA) 0.01000 Methyl-4-hydroxybenzoate 0.12500 Ethyl-4-hydroxybenzoate0.12500 Benzyl alcohol 0.26250 Polysorbate 80 0.42500 Palmitoyl ascorbicacid 0.09600 Tocopheryl linoleate 0.10000 +/− alpha tocopherol 0.05000Linalool 0.05000 Organic Phase Sub-Total 6.25800 Aqueous Phase -PAA/Tocopheryl Linoleate Intermediate Citric acid monohydrate 0.03600Disodium EDTA 0.05000 Sodium metabisulphite 0.02500 Disodium hydrogenorthophosphate 0.14400 dodecahydrate Water 22.65350 Aqueous PhaseSub-Total 22.90850 PAA/Tocopheryl Linoleate Tethersome 29.16650Intermediate Total Sum of Sequessome and 58.33300 TethersomeIntermediates Gel Phase - Final Product Sodium hydroxide 0.16000Carbopol 974P NF 0.75000 Glycerol 3.00000 Citric acid monohydrate0.05600 Disodium hydrogen orthophosphate 0.24200 dodecahydrate Water37.45900 Gel Phase Sub-Total 41.66700 Overall Total 100.00000

Summary Formulae of CBL-DERM-15-014

Ingredient (percentage; g per 100 g) Uneven Skin Tone ** SPC (dry mass)6.870 Polysorbate 80 0.850 Benzylalcohol 0.525 Methyl-4-hydroxybenzoate0.250 Ethyl-4-hydroxybenzoate 0.250 Butylhydroxyanisole 0.020 Linalool0.100 Sodium metabisulphite 0.050 Disodium EDTA 0.100 Disodium hydrogenphosphate 12 H₂O 0.530 Citric acid monohydrate 0.128 Glycerol 3.000Ethanol *** 3.255 Sodium hydroxide 0.160 Carbopol 974P NF 0.750 Water82.766  Agent of interest - tethered to Tethersomes Palmitoyl ascorbate0.096 Tocopheryl Linoleate 0.100 Tridecyl Salicylate 0.100 Agent ofinterest - in continuous phase Tocopherol 0.100 pH 5.5 *  * Final pHapproximate; to be confirmed ** The source of SC dry mass for UnevenSkin Tone will be lipoid Phospholipon 90K (P90K).

The quantity of P90K and ethanol to be added should be calculatedaccordingly.

Results

The product performed statistically favourably over the 6 week study inthe majority of attributes under Clearcast guidelines of advertising.The product did show a preference or favourability in the majority ofattributes:

-   -   After using the product, 100% of users agreed, or strongly        agreed the product reduced the appearance of fine lines and        wrinkles.    -   After using the product, 100% of users agreed, or strongly        agreed their skin looked significantly smoother.    -   After using the product, 100% of users agreed, or strongly        agreed their skin felt significantly more healthy.    -   After using the product, 96.36% of users agreed, or strongly        agreed their skin felt more elastic.    -   After using the product, 99.09% of users agreed, or strongly        agreed their complexion looks younger.    -   After using the product, 99.09% of users agreed, or strongly        agreed their complexion is visibly improved.    -   After using the product, 100% of users agreed, or strongly        agreed their complexion was visibly more radiant.    -   After using the product, 100% of users agreed, or strongly        agreed their complexion looked significantly healthier.    -   After using the product, 98.18% of users agreed, or strongly        agreed their skin tone looked significantly more even.    -   After using the product, 96.36% of users agreed, or strongly        agreed that the amount of hyperpigmentation/pigmented skin        blemishes were significantly reduced.    -   After using the product, 97.27% of users agreed, or strongly        agreed that the size of hyperpigmentation/pigmented skin        blemishes were significantly reduced.    -   After using the product, 99.09% of users agreed, or strongly        agreed that hyperpigmentation/pigmented skin blemishes were        significantly lighter.    -   After using the product, 20.00% of users agreed, or strongly        agreed that hyperpigmented/pigmented skin blemishes totally        disappeared.    -   After using the product, 100% of users agreed, or strongly        agreed their periorbital dark circles got significantly lighter.    -   After using the product, 95.45% of users agreed, or strongly        agreed this was the most effective hyperpigmentation solution        they had used.    -   After using the product, 92.73% of users stated, yes, they would        buy this product instead of their usual product.    -   After using the product, 100% of users stated, yes, they would        recommend this product to a friend.

With reference to packaging/promotional claims, any results with a top 3& 4 scoring greater than 80% are highly favourable.

1. A formulation comprising a first colloidal dispersion and a secondcolloidal dispersion, wherein each colloidal dispersion comprisesdeformable colloidal particles comprising a surfactant and optionally aphospholipid, and wherein one or more of the deformable colloidalparticles of the first colloidal dispersion comprise a modifiedcomponent comprising a first agent of interest (AOI), wherein themodified component is a lipid or surfactant bonded to an AOI, such thatthe entire AOI is outside the particle membrane, and wherein the firstAOI is zinc.
 2. The formulation of claim 1, wherein one or more of thedeformable colloidal particles of the second colloidal dispersioncomprise a second AOI and wherein the first AOI and the second AOI aredifferent.
 3. The formulation of claim 1, wherein one or more of thedeformable colloidal particles of the first colloidal dispersion and/orone or more of the deformable colloidal particles of the secondcolloidal dispersion comprise one or more further AOIs and wherein eachAOI is different.
 4. The formulation of claim 1, wherein the formulationcomprises one or more further colloidal dispersions comprisingdeformable colloidal particles comprising a surfactant and optionally aphospholipid and wherein one or more of the particles of at least two ofthe dispersions comprise one or more AOIs and wherein the first AOI,second AOI and each further AOI is different.
 5. through
 9. (canceled)10. The formulation of claim 1, comprising both a modified surfactantcomponent and a modified lipid component, optionally wherein the bondbetween each AOI and the lipid or surfactant of each modified componentis a covalent bond.
 11. (canceled)
 12. The formulation of claim 2,wherein the second AOI is selected from the group consisting of anelement, an ion, an inorganic salt, a small molecule, an amino acid, apeptide, a protein, a carbohydrate, a lipid, a micronutrient, amacromolecule or a macrocyclic molecule.
 13. The formulation of claim 2,wherein the second AOI is selected from the group consisting of zinc,ascorbate, tetrapeptide, tripeptide, salicylic acid, Vitamin D,tocopherol or menthol.
 14. (canceled)
 15. The formulation of claim 1,wherein the formulation comprises chlorhexidine, salicylic acid,capsaicin, caffeine or tocopherol, optionally wherein the chlorhexidine,salicylic acid, capsaicin, caffeine or tocopherol is not associated withthe deformable colloidal particles.
 16. (canceled)
 17. The formulationof claim 1, wherein the formulation comprises at least three colloidaldispersions.
 18. The formulation of claim 1, wherein the formulationcomprises at least four colloidal dispersions.
 19. The formulation ofclaim 1, wherein the formulation is topically applied.
 20. Theformulation of claim 1, wherein the formulation is applied weekly,between weekly and daily, daily, twice daily, three times a day or fourtimes a day.
 21. The formulation of claim 1, wherein the lipid isphosphatidylcholine.
 22. The formulation of claim 1, wherein thesurfactant is polysorbate
 80. 23. The formulation of claim 1, whereinthe formulation further includes one or more buffers, chelators,humectants, lubricants, antioxidants, preservatives, microbicides,antimicrobials, emollients, co-solvents or thickeners.
 24. through 36.(canceled)
 37. A method of cosmetically improving the appearance of asubject, the method comprising topically applying the formulation ofclaim 1 to the skin of a subject.
 38. through
 40. (canceled)
 41. Amethod of making the formulation of claim
 1. 42. A method of treating orpreventing a disease in a subject comprising topically applying theformulation of claim 1 to the subject, wherein the disease is treated,optionally wherein the disease is acne, dry skin, itchy skin, red skin,irritated skin, scaly skin, ageing skin, thinning skin, uneven skintone, periorbital skin, photo-aged skin or dermatitis.
 43. (canceled)44. A method of delivering an AOI to or through the skin of a patient,the method comprising topically applying to the skin of the patient theformulation of claim 1 in an amount sufficient to penetrate the skin todeliver the AOI.
 45. A kit comprising one or more compartments, whereinat least one compartment contains the formulation of claim 1, whereinthe kit comprises instructions for administrating the formulation to asubject in need thereof.
 46. (canceled)
 47. (canceled)